APOPTOSIS IN HEALTH AND DISEASE

1.0 INTRODUCTION

The study of programmed cell death, or apoptosis, has emerged from relative obscurity to become a major focus of research interest  in  many areas of medicine in the last decade. The driving force behind this attention has been a gradual recognition of the fundamental role played by apoptosis in normal development and tissue physiology, as well as in a surprisingly diverse collection of genetic and acquired diseases. Particularly significant advances have been made in defining the mechanisms of apoptotic control underlying the pathophysiology of viral infections, autoimmune diseases, neurodegenerative disorders, immunologic deficiencies, and cancers. Induction of apoptosis by the human immunodeficiency virus (HIV) in infected and uninfected cells appears to be integral to the pathophysiology of both the profound immunologic dysfunction and the dementia of AIDS (Gougeon and Montagnier 1993). Conversely, inhibition of apoptosis is critical to efficient replication and establishment of latency in many pathogenic viruses, most notably the Epstein-Barr virus associated with infectious mononucleosis, nasopharyngeal carcinoma, post-trans- plant lymphoproliferative disorder, and Burkitt’s lymphoma. Specific muta- tions of genes critical for apoptosis have been found in several autoimmune strains of mice and have been associated with autoimmune diseases in humans, including systemic lupus erythematosis (Mountz et al., 1994).  Programmed  cell death accounts for the necessary elimination of over 50% of neuronal cells in the developing brain, and aberrant control of apoptosis has been implicated not only in neurodegenerative disorders such as Alzheimer’s, Huntington’s, and Parkinson’s diseases, but also in various neurodevelopmental disorders includ- ing autism, Fragile X syndrome, and schizophrenia ( Margolis and  Chuang  1994). In cancer biology, alteration in the regulation of tumor cell survival is of critical impor- tance in the etiology and growth of tumors; it also provides clinically relevant prognostic information and will influence therapeutic decisions. The pharma- cology of almost all antineoplastic agents is much more strongly tied to induc- tion of apoptosis than had been imagined (Harrington et al., 1994 and Fisher 1994). Study of these diseases has brought into sharper focus, and in some cases redefined, the molecular mechanisms by which a cell regulates its survival and the signalling pathways that activate (or inhibit) this central apoptotic pathway. This review focuses on a number of recent, exciting molecular developments in this field

CHAPTER TWO

2.0 DISCOVERY AND ETYMOLOGY

German scientist Carl Vogt was first to describe the principle of apoptosis in 1842. In 1885, anatomist Walther Flemming delivered a more precise description of the process of programmed cell death. However, it was not until 1965 that the topic was resurrected. While studying tissues using electron microscopy, John Foxton Ross Kerr at University of Queensland was able to distinguish apoptosis from traumatic cell death (Kerr, 1965). Following the publication of a paper describing the phenomenon, Kerr was invited to join Alastair R Currie, as well as Andrew Wyllie, who was Currie's graduate student at University of Aberdeen. In 1972, the trio published a seminal article in the British Journal of Cancer. (Kerr et al., 1972). Kerr had initially used the term programmed cell necrosis, but in the article, the process of natural cell death was called apoptosis. Kerr, Wyllie and Currie credited James Cormack, a professor of Greek language at University of Aberdeen, with suggesting the term apoptosis. Kerr received the Paul Ehrlich and Ludwig Darmstaedter Prize on March 14, 2000, for his description of apoptosis. He shared the prize with Boston biologist Robert Horvitz. (O'Rourke and Ellem 2000). The 2002 Nobel Prize in Medicine was awarded to Sydney Brenner, Horvitz and John E. Sulston for their work identifying genes that control apoptosis. The genes were identified by studies in the nematode C. elegans and these same genes function in humans for apoptosis. John E. Sulston won the Nobel Prize in Medicine in 2002, for his pioneering research on apoptosis.

In Greek, apoptosis translates to the "dropping off" of petals or leaves from plants or trees. Cormack, professor of Greek language, reintroduced the term for medical use as it had a medical meaning for the Greeks over two thousand years before. Hippocrates used the term to mean "the falling off of the bones". Galen extended its meaning to "the dropping of the scabs". Cormack was no doubt aware of this usage when he suggested the name. Debate continues over the correct pronunciation, with opinion divided between pronunciations with the second p silent second p pronounced as in the original Greek. (Kerr Wyllie and Currie paper 1972) In English, the p of the Greek -pt-consonant cluster is typically silent at the beginning of a word (e.g. pterodactyl, Ptolemy), but articulated when used in combining forms preceded by a vowel, as in helicopter or the orders of insects: diptera,  lepidoptera, etc.

In the original Kerr Wyllie and Currie paper, British Journal of Cancer, 1972, there is a footnote regarding the pronunciation:

"We are most grateful to Professor James Cormack of the Department of Greek, University of Aberdeen, for suggesting this term. The word "apoptosis"  is used in Greek to describe the "dropping off" or "falling off" of petals from flowers, or leaves from trees. To show the derivation clearly, we propose that the stress should be on the penultimate syllable, the second half of the word being pronounced like "ptosis" (with the "p" silent), which comes from the same root "to fall", and is already used to describe the drooping of the upper eyelid."

2.1 CAUSES OF APOPTOSIS

Causes of apoptosis and necrosis cell death

Cell death is completely normal and it goes on all the time in your body. In fact, death of cells is essential for healthy development and homeostasis. However, not all cell death is beneficial.

Beneficial cell death is one that is carried out in a controlled manner. Whenever a single cell malfunctions or becomes diseased, then apoptosis occurs. This is a highly regulated breakdown of that cell coupled with the production of a new one. Thus, maintaining a relatively constant process of life and death. 

Some causes for apoptosis to occur include:

·         cells infected with a virus

·         cells impaired beyond repair

·         cells in a stressful state  starvation

·         cell DNA damaged by oxidants or other agents

Apoptosis is an organized way of removing a non-functioning cell so that it no longer drains nutrients nor spreads infection.

On the other hand, necrosis means death of a group cells in the same area caused by the lack of blood. Necrosis caused demise is ungoverned and irreversible

What also causes these morphological changes that we recognize as apoptosis and the biochemical changes often associated with this phenomenon? The answer is proteases. Specifically, activation of a family of intracellular cysteine proteases which cleave their substrates at aspartic acid residues, known as caspases for Cysteine Aspartyl-specific Proteases. These proteases are present as inactive zymogens in essentially all animal cells, but can be triggered to assume active states, generally involving their proteolytic processing at conserved aspartic acid (Asp) residues. During activation, the zymogen pro-proteins are cleaved to generate the large (∼20 kd) and small (∼10 kd) subunits of the active enzymes, typically liberating an N-terminal prodomain from the processed polypeptide chain. The active enzymes consist of heterotetramers composed of two large and two small subunits, with two active sites per molecule. Analysis of the structures of the active sites of these enzymes, experiments with combinatorial peptide libraries, and other data suggest that caspases recognize the Asp residues they cleave within the context of tetrapeptide motifs, where the most proximal (N-terminal) residue recognized is designated P4 (position 4) and target Asp is P1 (position 1), and where cleavage occurs at the peptidyl bond distal (C-terminal) to the targeted Asp. This information about the structures and mechanisms of caspases has been exploited for developing small-molecule inhibitors, which are finding their way into clinical trials for stroke, liver failure, inflammatory diseases, and a wide variety of other ailments.

The observation that caspases cleave their substrates at Asp residues and are also activated by proteolytic processing at Asp residues makes evident that these proteases collaborate in proteolytic cascades, whereby caspases activate themselves and each other. In humans and mice, approximately 14 caspases have been identified. They can be subgrouped according to either their amino acid sequence similarities or their protease specificities. From a functional perceptive, it is useful to view the caspases as either upstream (initiator) caspases or downstream (effector) caspases. The proforms of upstream initiator caspases possess large N-terminal pro-domains, which function as protein interaction modules, allowing them to interact with various proteins that trigger caspase activation. In contrast, the proforms of downstream effector caspases contain only short N-terminal prodomains, serving no apparent function. Downstream caspases are largely dependent on upstream caspases for their proteolytic processing and activation. Accordingly, the sequence of the cleavage sites separating the large and small subunits of the zymogen forms of the effector caspases generally match the preferred tetrapeptide specificities of the upstream initiator caspases. Similarly, examination of the cleavage sites of multiple cellular proteins, which have been identified as caspase substrates and which are known to undergo processing during apoptosis, reveals (in most instances) coincidence with the preferred tetrapeptide sequences cleaved by the effector caspases. These substrates of effector caspases include protein kinases (often separating the autorepressing regulatory domains from catalytic domains) and other signal transduction proteins, cytoskeletal and nuclear matrix proteins, chromatin-modifying (eg, polyADP ribosyl polymerase) and DNA repair proteins, and inhibitory subunits of endonucleases (CIDE family proteins).

Though most caspases are directly involved in cell death, a few are not, at least in mammals and higher eukaryotes. A subgroup of caspases, including caspase-1,- 4, and -5 in humans, is involved in processing of pro-inflammatory cytokines such as pro-interleukin-1β (pro-IL-1β) and pro-IL-18. Unlike the effector caspases, which induce apoptosis, the tetrapeptide specificities of these cytokine-processing proteases do not match the cleavage sites of most of the proteins known to undergo cleavage during apoptosis, but they do coincide with the sequences of the cleavage sites within pro-cytokines

2.4 Schematic representation of the main molecular pathways leading to apoptosis

Plate 2: Molecular pathways leading to apoptosis

Extrinsic apoptosis indicates a form of death induced by extracellular signals that result in the binding of ligands to specific trans-membrane receptors, collectively known as death receptors (DR) belonging to the TNF/NGF family. All death receptors function in a similar way: upon ligand binding several receptor molecules are brought together and undergo conformational changes allowing the assembly of a large multi-protein complex known as Death Initiation Signalling Complex (DISC) that leads to activation of the caspase cascade. In the FAS/CD95 signalling complex, that can be used as a prototype of this form of death, upon ligand binding FAS recruits, through a highly conserved 80 amino acid domain, known as death domain (DD), an adaptor molecule: Fas-associated protein with a DD (FADD). FADD contains another conserved protein interaction domain known as Death Effector Domain (DED) that binds to a homologous domain in caspase 8 leading to its activation. Active caspase 8 will activate additional caspase 8 molecules as well as downstream caspases such as caspase 3. (Lavrit and  Krammer.  2010)

The intrinsic pathway is activated in response to a number of stressing conditions including DNA damage, oxidative stress and many others. In all cases this multiple forms of stress converge on the mitochondria and determine mitochondrial outer membrane permeabilization (MOMP) this in turn results in dissipation of the mitochondrial membrane potential and therefore in cessation of ATP production as well as release of a number of proteins that contribute to caspase activation. At least two molecular mechanisms (not mutually exclusive) have been proposed to explain how different signals converge at the mitochondria resulting in MOMP. One involves the pore forming ability of some of the BCL-2 family proteins in the outer mitochondrial membrane and the other is the result of the opening in the inner membrane (Gavathiotis and walensky. 2011). of the permeability transition pore complex (PTPC) that would require the Adenine Nucleotide Transporter (ANT) and the Voltage Dependent Anion Channel (VDAC). (Gavathiotis and Walensky . 2011), (Yivgi-ohana et al., 2011).The Bcl-2 family proteins are essential regulators of this type of apoptosis and are all characterized by the presence of at least one Bcl-2 Homology (BH) domain. From a functional point of view they can be classified in anti-apoptotic members containing three or four BH domains (such as Bcl-2, Bcl-xl, Bcl-w, Mcl-1) and pro-apoptotic members with two or three BH domains (such as Bax, Bak, Bcl-xs, Bok) or with just one (such as Bad, Bik, Bid, Bim, Noxa, Puma). Pro-apoptotic members of the family mediate apoptosis by disrupting membrane integrity either directly forming pores or by binding to mitochondrial channel proteins such as VDAC or ANT, while anti-apoptotic members would prevent apoptosis by interfering with pro-apoptotic member aggregation. The different apoptotic signals are sensed by BH3 only proteins that are induced or activated and migrate to the mitochondria where they bind the pro-survival members of the family removing their block or to the pro-apoptotic members promoting their aggregation. (Chipuk et al., 2008)

In any case once MOMP occurs a number of proteins are released from the mitochondria, these include Cytochrome C (CYTC), apoptosis-inducing factor (AIF), endonuclease G (endo G), Direct IAP-binding protein with low PI (DIABLO, also known as SMAC) and others. Once CYTC is released it binds to APAF-1 inducing the formation of a large complex, known as the apoptosome that recruits caspase 9. In the apoptosome, caspase 9 is activated and cleaved and will activate additional molecules of caspase 9 as well as down-stream caspases such as caspase 3. Due to its lethality the system is subject to a number of controls as an example the cytoplasm contains a class of proteins known as Inhibitors of Apoptosis IAPS that bind and inactivate caspases. Upon MOMP the mitochondria also releases proteins such as DIABLO/SMAC that bind to IAPS removing their inhibition and allowing apoptosis to occur. (Kaufmann et al., 2012)

The intrinsic and extrinsic pathways are not completely independent: in some cells in fact activation of caspase 8 results in activation of the mitochondrial pathway. In this case caspase 8 among other things cleaves a BH3 only protein BID generating a truncated fragment known as truncated BID (tBID) that can permeabilize the mitochondrion resulting in MOMP (Kaufmann  et al., 2012) 

CHAPTER THREE

3.0 REMOVAL OF DEAD CELLS

The removal of dead cells by neighboring phagocytic cells has been termed efferocytosis. Dying cells that undergo the final stages of apoptosis display phagocytotic molecules, such as phosphatidylserine, on their cell surface Phosphatidylserine is normally found on the cytosolic surface of the plasma membrane, but is redistributed during apoptosis to the extracellular surface by a protein known as scramblase. These molecules mark the cell for phagocytosis by cells possessing the appropriate receptors, such as macrophages. Upon recognition, the phagocyte reorganizes its cytoskeleton for engulfment of the cell. The removal of dying cells by phagocytes occurs in an orderly manner without eliciting an inflammatory response (Gavathiotis and Walensk, 2011).

3.1 Disease Associated With Apoptosis

3.1.0 Neurological disorders

From a physiological point of view apoptosis plays a key role in central nervous development, while in adult brain it is involved in the pathogenesis of a number of diseases including neurodegenerative diseases and acute injury (i.e. stroke).

3.1.1 Neurodegenerative diseases

CHAPTER FOUR

4.1   MANAGEMENT OF THE DISEASES

4.1.0 Management of cancer

Cancer can be treated by surgery, chemotherapy, radiation therapy, immunotherapy, and monoclonal antibody therapy. The choice of therapy depends upon the location and grade of the tumor and the stage of the disease, as well as the general state of the patient (performance status). A number of experimental cancer treatments are also under development.

Complete removal of the cancer without damage to the rest of the body is the goal of treatment. Sometimes this can be accomplished by surgery, but the propensity of cancers to invade adjacent tissue or to spread to distant sites by microscopic metastasis often limits its effectiveness; chemotherapy and radiotherapy can unfortunately have a negative effect on normal cells. (Enge et al., 2012).

Because "cancer" refers to a class of diseases, it is unlikely that there will ever be a single "cure for cancer" any more than there will be a single treatment for all infectious diseases (Wanjek and Christopher, 2009). Angiogenesis inhibitors were once thought to have potential as a "silver bullet" treatment applicable to many types of cancer, but this has not been the case in practice (Hayden and Erika 2009).

4.1.1 Types of treatments

The treatment of cancer has undergone evolutionary changes as understanding of the underlying biological processes has increased. Tumor removal surgeries have been documented in ancient Egypt, hormone therapy was developed in 1896, and radiation therapy was developed in 1899. Chemotherapy, Immunotherapy, and newer targeted therapies are products of the 20th century. As new information about the biology of cancer emerges, treatments will be developed and modified to increase effectiveness, precision, survivability, and quality of life.

4.1.2 Surgery

In theory, non-hematological cancers can be cured if entirely removed by surgery, but this is not always possible. When the cancer has metastasized to other sites in the body prior to surgery, complete surgical excision is usually impossible. In the Halstedian model of cancer progression, tumors grow locally, and then spread to the lymph nodes, then to the rest of the body. This has given rise to the popularity of local-only treatments such as surgery for small cancers. Even small localized tumors are increasingly recognized as possessing metastatic potential.

Examples of surgical procedures for cancer include mastectomy for breast cancer, prostatectomy for prostate cancer, and lung cancer surgery for non-small cell lung cancer. The goal of the surgery can be either the removal of only the tumor, or the entire organ. A single cancer cell is invisible to the naked eye but can regrow into a new tumor, a process called recurrence. For this reason, the pathologist will examine the surgical specimen to determine if a margin of healthy tissue is present, thus decreasing the chance that microscopic cancer cells are left in the patient.

In addition to removal of the primary tumor, surgery is often necessary for staging, e.g. determining the extent of the disease and whether it has metastasized to regional lymph nodes. Staging is a major determinant of prognosis and of the need for adjuvant therapy.

Occasionally, surgery is necessary to control symptoms, such as spinal cord compression or bowel obstruction. This is referred to as palliative treatment.

If surgery is possible and appropriate, it is commonly performed before other forms of treatment, although the order does not affect the outcome. (Kolata and Gina 2009) In some instances, surgery must be delayed until other treatments are able to shrink the tumor.

4.1.3 Radiation therapy

Radiation therapy (also called radiotherapy, X-ray therapy, or irradiation) is the use of ionizing radiation to kill cancer cells and shrink tumors. Radiation therapy can be administered externally via external beam radiotherapy (EBRT) or internally via brachytherapy. The effects of radiation therapy are localized and confined to the region being treated. Radiation therapy injures or destroys cells in the area being treated (the "target tissue") by damaging their genetic material, making it impossible for these cells to continue to grow and divide. Although radiation damages both cancer cells and normal cells, most normal cells can recover from the effects of radiation and function properly. The goal of radiation therapy is to damage as many cancer cells as possible, while limiting harm to nearby healthy tissue. Hence, it is given in many fractions, allowing healthy tissue to recover between fractions.

Radiation therapy may be used to treat almost every type of solid tumor, including cancers of the brain, breast, cervix, larynx, liver, lung, pancreas, prostate, skin, stomach, uterus, or soft tissue sarcomas. Radiation is also used to treat leukemia and lymphoma. Radiation dose to each site depends on a number of factors, including the radio sensitivity of each cancer type and whether there are tissues and organs nearby that may be damaged by radiation. Thus, as with every form of treatment, radiation therapy is not without its side effects.

4.1.4 Chemotherapy

Chemotherapy is the treatment of cancer with drugs ("anticancer drugs") that can destroy cancer cells. In current usage, the term "chemotherapy" usually refers to cytotoxic drugs which affect rapidly dividing cells in general, in contrast with targeted therapy. Chemotherapy drugs interfere with cell division in various possible ways, e.g. with the duplication of DNA or the separation of newly formed chromosomes. Most forms of chemotherapy target all rapidly dividing cells and are not specific to cancer cells, although some degree of specificity may come from the inability of many cancer cells to repair DNA damage, while normal cells generally can. Hence, chemotherapy has the potential to harm healthy tissue, especially those tissues that have a high replacement rate (e.g. intestinal lining). These cells usually repair themselves after chemotherapy.

Because some drugs work better together than alone, two or more drugs are often given at the same time. This is called "combination chemotherapy"; most chemotherapy regimens are given in a combination (Takimoto et al., 2008)

The treatment of some leukaemias and lymphomas requires the use of high-dose chemotherapy, and total body irradiation (TBI). This treatment ablates the bone marrow, and hence the body's ability to recover and repopulate the blood. For this reason, bone marrow, or peripheral blood stem cell harvesting is carried out before the ablative part of the therapy, to enable "rescue" after the treatment has been given. This is known as autologous stem cell transplantation. Alternatively, hematopoietic stem cells may be transplanted from a matched unrelated donor (MUD).

4.2.0 MANAGEMENT OF PARKINSON'S DISEASE

Treatment for Parkinson's disease (PD), due to its chronic nature, requires broad-based management including patient and family education, support group services, general wellness maintenance, exercise, and nutrition. At present, there is no cure for PD, but medications or surgery can provide relief from the symptoms.

While many medications treat Parkinson's, none actually reverse the effects of the disease or cure it. Furthermore, the gold standard treatment varies with the disease state. People with Parkinson's therefore often must take a variety of medications to manage the disease's symptoms. Several medications currently in development seek to better address motor fluctuations and nonmotor symptoms of PD. However, none are yet on the market with specific approval to treat Parkinson's. (Bronstein et al., 2010)

4.2.1 Pharmacologic

The main families of drugs useful for treating motor symptoms are Levodopa, dopamine agonists and MAO-B inhibitors (The National Collaborating Centre for Chronic Conditions, ed. 2006). The most commonly used treatment approach varies depending on the disease stage. Two phases are usually distinguished: an initial phase in which the individual with PD has already developed some disability for which he needs pharmacological treatment, and a second stage in which the patient develops motor complications related to levodopa usage (The National Collaborating Centre for Chronic Conditions, ed. 2006). Treatment in the initial state aims to attain an optimal tradeoff between good management of symptoms and side-effects resulting from enhancement of dopaminergic function. The start of L-DOPA treatment may be delayed by using other medications such as MAO-B inhibitors and dopamine agonists, in the hope of causing the onset of dyskinesia’s to be retarded (The National Collaborating Centre for Chronic Conditions, ed. 2006). In the second stage the aim is to reduce symptoms while controlling fluctuations of the response to medication. Sudden withdrawals from medication, and overuse by some patients, also have to be controlled. When medications are not enough to control symptoms, surgical techniques such as deep brain stimulation can relieve the associated movement disorders (Alterman et al., 2010).

4.2.2 Levodopa

Plate: Drugs used for the treatment of Parkinson's disease

Stalevo, a commercial preparation combining entacapone, levodopa and carbidopa for treatment of Parkinson's disease

Plate 4: Circuits of the basal ganglia in treatment of Parkinson's disease.

Circuits of the basal ganglia in treatment of Parkinson's disease. Model of the effect of medication on motor symptoms: levodopa, dopamine agonists and MAO-B inhibitors stimulate excitatory signals from the thalamus to the cortex by effects on the striatum, compensating for decreased dopaminergic signals from substantia nigra.

Levodopa (or L-DOPA) has been the most widely used treatment for over 30 years. L-DOPA is transformed into dopamine in the dopaminergic neurons by dopa-decarboxylase. Since motor symptoms are produced by a lack of dopamine in the substantia nigra the administration of L-DOPA temporarily diminishes the motor symptomatology.

Only 5-10% of L-DOPA crosses the blood–brain barrier. The remaining is often metabolised to dopamine elsewhere, causing a wide variety of side effects including nausea, dyskinesias and stiffness (Frosini et al., 2009). Carbidopa and benserazide are peripheral dopa decarboxylase inhibitors. They inhibit the metabolism of L-DOPA in the periphery thereby increasing levodopa delivery to the central nervous system. They are generally given as combination preparations with levodopa. Existing preparations are carbidopa/levodopa (co-careldopa, trade names Sinemet, Parcopa, Atamet) and benserazide/levodopa (co-beneldopa, trade name Madopar). Levodopa has also been related to a dopamine dysregulation syndrome, which is a compulsive overuse of the medication, and punding (Ceravolo, et al., 2009).

There are controlled release versions of Sinemet and Madopar that spread out the effect of the levodopa. Duodopa is a combination of levodopa and carbidopa. Slow-release levodopa preparations have not shown an increased control of motor symptoms or motor complications when compared to immediate release preparations.

Tolcapone inhibits the catechol-O-methyltransferase COMT enzyme, which degrades dopamine and levadopa, thereby prolonging the therapeutic effects of levodopa. It, alongside inhibitors of peripheral dopa decarboxylase, have been used to complement levodopa. However, due to its possible side effects such as liver failure, it's limited in its availability. A similar drug, entacapone has not been shown to cause significant alterations of liver function and maintains adequate inhibition of COMT over time. Entacapone is available for treatment alone (COMTan) or combined with carbidopa and levodopa (Stalevo).

Levodopa results in a reduction in the endogenous formation of L-DOPA, and eventually becomes counterproductive. Levodopa preparations lead in the long term to the development of motor complications characterized by involuntary movements called dyskinesias and fluctuations in the response to medication. When this occurs PD patients change rapidly from stages with good response to medication and few symptoms ("on" state) to phases with no response to medication and important motor symptoms ("off" state). For this reason levodopa doses are kept as low as possible while maintaining functionality. Delaying the initiation of dopatherapy, using instead alternatives for some time, is also common practice. A former strategy to reduce motor complications was to withdraw patients from L-DOPA for some time. It is discouraged now since it can bring dangerous side effects such as neuroleptic malignant syndrome. Most people will eventually need levodopa and later develop motor complications.

The on-off phenomenon is an almost invariable consequence of sustained levodopa treatment in patients with Parkinson's disease. Phases of immobility and incapacity associated with depression alternate with jubilant thaws. Both pharmacokinetic and pharmacodynamic factors are involved in its pathogenesis, but evidence is presented to indicate that the importance of levodopa handling has been underestimated and that progressive reduction in the storage capacity of surviving nigrostriatal dopamine terminals is not a critical factor. Re-distribution of levodopa dosage which may mean smaller, more frequent doses, or larger less frequent increments, may be helpful in controlling oscillations in some patients. Dietary protein restriction, the use of selegiline hydrochloride and bromocriptine may also temporarily improve motor fluctuations. New approaches to management include the use of subcutaneous apomorphine, controlled-release preparations of levodopa with a peripheral dopa decarboxylase inhibitor and the continuous intra-duodenal administration of levodopa.

4.2.3 Dopamine agonists

Dopamine agonists in the brain have a similar effect to levodopa since they bind to dopaminergic post-synaptic receptors. Dopamine agonists were initially used for patients experiencing on-off fluctuations and dyskinesias as a complementary therapy to levodopa but they are now mainly used on their own as an initial therapy for motor symptoms with the aim of delaying motor complications (Goldenberg 2008). When used in late PD they are useful at reducing the off periods. Dopamine agonists include bromocriptine, pergolide, pramipexole, ropinirole, piribedil, cabergoline, apomorphine, and lisuride.

Agonists produce significant, although mild, side effects including somnolence, hallucinations, insomnia, nausea, and constipation. Sometimes side effects appear even at a minimal clinically efficacious dose, leading the physician to search for a different agonist or kind of drug. When compared with levodopa, while they delay motor complications they control worse symptoms. Nevertheless they are usually effective enough to manage symptoms in the initial years. They are also more expensive (Samii, Nutt and Ransom 2004). Dyskinesias with dopamine agonists are rare in younger patients, but along other side affects more common in older patients (Samii et al., 2004). All this has led to agonists being the preferential initial treatment for the former as opposed to levodopa in the latter. Agonists at higher doses have also been related to a wide variety of impulse control disorders (Ceravolo, Frosini, Rossi and Bonuccelli 2009).

Apomorphine, which is a non-orally administered dopamine agonist, may be used to reduce off periods and dyskinesia in late PD. Since secondary effects such as confusion and hallucinations are not rare it has been recommended that patients under apomorphine treatment should be closely monitored. Apomorphine can be administered via subcutaneous injection using a small pump which is carried by the patient. A low dose is automatically administered throughout the day, reducing the fluctuations of motor symptoms by providing a steady dose of dopaminergic stimulation. After an initial "apomorphine challenge" in hospital to test its effectiveness and brief patient and primary caregiver (often a spouse or partner), the latter of whom takes over maintenance of the pump. The injection site must be changed daily and rotated around the body to avoid the formation of nodules. Apomorphine is also available in a more acute dose as an auto injector pen for emergency doses such as after a fall or first thing in the morning. Nausea and vomiting are common, and may require domperidone (an antiemetic).

4.2.4 MAO-B inhibitors

MAO-B inhibitors (Selegiline and rasagiline) increase the level of dopamine in the basal ganglia by blocking its metabolization. They inhibit monoamine oxidase-B (MAO-B) which breaks down dopamine secreted by the dopaminergic neurons.Therefore reducing MAO-B results in higher quantities of L-DOPA in the striatum. Similarly to dopamine agonists, MAO-B inhibitors improve motor symptoms and delay the need of taking levodopa when used as monotherapy in the first stages of the disease but produce more adverse effects and are less effective than levodopa. Evidence on their efficacy in the advanced stage is reduced although it points towards them being useful to reduce fluctuations between on and off periods. Although an initial study had as result that selegiline in combination with levodopa increased the risk of death this has been later disproven.

Metabolites of selegiline include L-amphetamine and L-methamphetamine (not to be confused with the more notorious and potent dextrorotary isomers). This might result in side effects such as insomnia. Another side effect of the combination can be stomatitis. Unlike other non-selective monoamine oxidase inhibitors, tyramine-containing foods do not cause a hypertensive crisis.

4.2.5 Other Drugs

There is some indication that other drugs such as amantadine and anticholinergics may be useful as treatment of motor symptoms in early and late PD, but since quality of evidence on efficacy is reduced they are not first choice treatments. In addition to motor PD is accompanied by an ample range of different symptoms. Different compounds are used to improve some of these problems (Hasnain et al., 2009). Examples are the use of clozapine for psychosis, cholinesterase inhibitors for dementia and modafinil for day somnolence (Hasnain et al., 2009).

A preliminary study indicates that taking the drug donepezil (Aricept) may help prevent falls in people with Parkinson's. Donepezil boosts levels of the neurotransmitter acetylcholine, and is currently an approved therapy for the cognitive symptoms of Alzheimer's disease (Parkinson's disease Foundation Science News. 2010). In the study, participants taking donepezil experienced falls half as often as those taking a placebo, and those who previously fell the most showed the most improvement (Chung, Lobb, Nutt, Horak, 2010).

The introduction of clozapine (Clozaril) represents a breakthrough in the treatment of psychotic symptoms of PD. Prior to its introduction, treatment of psychotic symptoms relied on reduction of dopamine therapy or treatment with first generation antipsychotics, all of which worsened motor function. Other atypical antipsychotics useful in treatment include quetiapine (Seroquel), ziprasidone (Geodon), aripiprazole (Abilify), and paliperidone (Invega). Clozapine is believed to have the highest efficacy and lowest risk of extrapyramidal side effect. (Hasnain et al., 2009)

 4.2.6 Surgical

Plate 5: Illustration showing an electrode placed deep seated in the brain

Treating PD with surgery was once a common practice. But after the discovery of levodopa, surgery was restricted to only a few cases (The National Collaborating Centre for Chronic Conditions, ed. 2006). Studies in the past few decades have led to great improvements in surgical techniques, and surgery is again being used in people with advanced PD for whom drug therapy is no longer sufficient. (The National Collaborating Centre for Chronic Conditions, ed. 2006).

Less than 10% of PD sufferers qualify as suitable candidates for a surgical response. There are three different mechanisms of surgical response for PD: ablative surgery, (the irreversible burning or freezing of brain tissue) stimulation surgery or deep brain stimulation (DBS), and transplantation or restorative surgery (Parkinson's disease surgery neurology Channel. Retrieved on 2010). Target areas for DBS or lesions include the thalamus, the globus pallidus (the lesion technique being called pallidotomy) or the subthalamic nucleus.

4.2.7 Neuroablative Lesion Surgery

Neuroablative Lesion surgery (NAS) locates and destroys, by heat, the parts of the brain that are associated with producing Parkinsonian neurological symptoms. The procedures generally involve a thalamotomy and/or pallidotomy.

A thalamotomy is the destruction of a part of the thalamus, in particular the ventralis intermedius, in order to suppress tremor in 80-90% of patients. If rigidity and akinesia are apparent, the subthalamis nucleus is then the site of ablation.

A pallidotomy involves the destruction of the globus pallidus, in particular the globus pallidus interna, in patients with Parkinson's that suffer from rigidity and akinesia.

Because it is difficult to accurately measure the amount of tissue to be destroyed, it is not uncommon for tremors to persist through multiple courses of surgery since tissue is irreversibly damaged and removed and it is safer to test smaller areas of tissue to prevent serious complications, such as a stroke This method has been generally replaced by deep brain surgery.

4.2.8 Deep Brain Stimulation

Deep brain stimulation (DBS) is presently the most used method of surgical treatment because it does not destroy brain tissue, it is reversible, and it can be tailored to each individual at their particular stage of disease. DBS employs three hardware components: a neurostimulator, also called an implanted pulse generator (IPG), which generates electrical impulses used to modulate neural activity, a lead wire which directs the impulses to a number of metallic electrodes towards the tip of the lead near the stimulation target, and an extension wire that connects the lead to the IPG. The IPG, which is battery-powered and encased in titanium, is traditionally implanted under the collarbone, and is connected by the subcutaneous extension to the lead, which extends from outside the skull under the scalp down into the brain to the target of stimulation. The entire three component system is sometimes referred to as a brain pacemaker, as the system operates on many of the same principles as medical cardiac pacing.

The pre-operative targeting of proper implantation sites can be accomplished via the indirect and direct methods.

The indirect method utilizes computer tomography, magnetic resonance imaging, or ventriculography to locate the anterior and posterior commissures and then employs pre-determined coordinates and distances from the intercommissural line in order to define the target area. Subsequent histologically defined atlas maps can also be used to verify the target area (Nolte, 2012). The direct method provides visualization and targeting of deep nuclei by applying stereotactic pre-operative MRI, which unlike the indirect method, takes into account the anatomic variation of the nuclei’s size, position, and functional segregation amongst individuals (Nolte, 2012).

Electrophysial functional mapping (EFM), a tool utilized in both methods in order to verify the target nuclei, has come under scrutiny due to its associated risks of hemorrhages, dysarthria or tetanic contractions. Recently, Susceptibility Weighted Imaging (SWI), a type of MRI has shown incredible resolving power in its ability to distinguish these deep brain nuclei and is being used in DBS in order to reduce the over-use of EFM (Abosch, 2010).

DBS is recommended to PD patients without important neuropsychiatric contraindications who suffer motor fluctuations and tremor badly controlled by medication, or to those who are intolerant to medication (Bronstein et al., 2010). DBS is effective in suppressing symptoms of PD, especially tremor. A recent clinical study led to recommendations on identifying which Parkinson's patients are most likely to benefit from DBS (Bronstein et al., 2010).

4.2.9 DIET

Muscles and nerves that control the digestive process may be affected by PD, therefore, it is common to experience constipation and gastroparesis (food remaining in the stomach for a longer period of time than normal) (Barichella et al., 2009). A balanced diet is recommended to help improve digestion. Diet should include high-fiber foods and plenty of water. Levodopa and proteins use the same transportation system in the intestine and the blood–brain barrier, competing between them for access. When taken together the consequences of such competition is a reduced effectiveness of the drug. Therefore when levodopa is introduced excessive proteins are discouraged, while in advanced stages additional intake of low-protein products such as bread or pasta is recommended for similar reasons. To minimize interaction with proteins levodopa is recommended to be taken 30 minutes before meals. At the same time, regimens for PD restrict proteins during breakfast and lunch and are usually taken at dinner. As the disease advances dysphagia may appear. In such cases specific measures include the use of thickening agents for liquid intake, special postures when eating and gastrostomy in the worst cases (Barichella et al., 2009).

4.2.10 Rehabilitation

There is partial evidence that speech or mobility problems can improve with rehabilitation although studies are scarce and of low quality (Goodwin et al., 2008) Regular physical exercise and/or therapy can be beneficial to maintain and improve mobility, flexibility, strength, gait speed, and quality of life. Exercise may also improve constipation. Exercise interventions have been shown to benefit patients with Parkinson’s disease in regards to physical functioning, health-related quality of life, and balance and fall risk. In a review of 14 studies examining the effects of exercise on persons with Parkinson’s disease, no adverse events or side-effects occurred following any of the exercise interventions (Goodwin et al., 2008). There are five proposed mechanisms by which exercise enhances neuroplasticity. Intensive activity maximizes synaptic plasticity; 2) complex activities promote greater structural adaptation; 3) activities that are rewarding increase dopamine levels and therefore promote learning/relearning; 4) dopaminergic neurones are highly responsive to exercise and inactivity (“use it or lose it 5) where exercise is introduced at an early stage of the disease, progression can be slowed. (Ramig et al., 2006). One of the most widely practiced treatment for speech disorders associated with Parkinson's disease is the Lee Silverman Voice Treatment (LSVT), which focuses on increasing vocal loudness and has an intensive approach of one month (Fox et al., 2006).Speech therapy and specifically LSVT may improve voice and speech function. Occupational therapy (OT) aims to promote health and quality of life by helping people with the disease to participate in as many activities of their daily living as possible. There have been few studies on the effectiveness of OT and their quality is poor, although there is some indication that it may improve motor skills and quality of life for the duration of the therapy (Dixon et al., 2007).

4.2.10 Palliative care

Palliative care is often required in the final stages of the disease, often when dopaminergic treatments have become ineffective. The aim of palliative care is to achieve the maximum quality of life for the person with the disease and those surrounding him or her. Some central issues of palliative are; caring for patients at home while adequate care can be given there; reducing or withdrawing dopaminergic drug intake to reduce drug side effects and complications; preventing pressure ulcers by management of pressure areas of inactive patients; facilitating the patient's end of life decisions for the patient as well as involved friends and relatives (The National Collaborating Centre for Chronic Conditions, ed. 2006).  

 CHAPTER FIVE

CONCLUSIONS

A number of disparate signals are capable of modulating apoptosis in different cells and in different contexts, and these signals appear to focus in on a central regulatory pathway–determining cell fate. Many of these signalling pathways, including components of the central apoptotic pathway, are evolutionarily conserved. The pathways that have been outlined have yielded important in- sights into organism development, tissue homeostasis, and the pathophysiology of whole categories of disease. Major gaps in our understanding of the regulation of apoptosis include the chain of events connecting ligand binding to cell surface receptors with the central apoptotic pathway, the mechanisms of action of the Bcl-2– and ICE-related proteins, the communication between Bcl-2 and ICE family members, and the mechanisms directly responsible for the characteristic cytoplasmic and nuclear changes of apoptosis. Clearly, greater definition of the essential connections among these various pathways is required. The ultimate challenge may be to translate the  knowledge  gained  into therapeutic strategies to improve clinical outcome in the many diseases linked to disregulation of apoptosis. In cancer research, the surprising finding that the cytotoxic effects of chemotherapeutic agents operate primarily through induc- tion of tumor cell apoptosis has prompted an investigation of anti-neoplastic therapies that more directly target the aberrant control of apoptosis in tumors. A pilot study of antisense bcl-2 oligonucleotides in the treatment of B cell lymphomas is currently being conducted (Friedman, et al., 1995.). Direct antitumor therapy target- ing apoptotic modulation may prove to be much less systemically toxic than standard chemotherapy and could also be used in an adjuvant manner, to increase the apoptotic susceptibility of tumours at the time they are exposed to chemotherapy. In the treatment of autoimmune disease, targeted induction of apoptosis in autoimmune subsets of lymphocytes may be possible using the specific autoantigen in the absence of costimulatory survival signals. The potential utility of this strategy was demonstrated in mice treated for experi- mental autoimmune encephalitis (Critchfield et al., 1994.). Targeted modulation of EBV LMP- 1–derived signaling may be an ideal way to specifically treat post-transplant lymphoproliferative disorder without risking transplant rejection and may play an important role in the management of EBV-positive lymphomas. Finally, modification of the apoptotic induction in uninfected lymphocytes in HIV infection, which has been linked to effects of free extracellular gp120 and Tat protein (Kornfield et al., 1988 Banda et al., 1992 Friedman et al., 1995) May have a major impact on the progression of AIDS. The era of widespread clinical implementation of apoptotic modulation in the treat- ment of disease has not yet arrived, but it has the potential for tremendous impact on the prognosis of many important and challenging diseases.

REFERENCES

Agency for Science, Technology and Research. "Prof Andrew H. Wyllie -  Lecture Abstract". Archived from the original on 2007-11-13. Retrieved  2007-03-3

Ai X, Butts B, Vora K, Li W, Tache-Talmadge C, Fridman A, Mehmet H. Generation and characterization of antibodies specific for caspase-cleaved neo-epitopes: a novel approach. Cell Death Dis. 2011;2:e205. 

 Alberts, Bruce; Johnson, Alexander; Lewis, Julian; Raff, Martin; Roberts, Keith; Walter, Peter 2008. "Chapter 18 Apoptosis: Programmed Cell  Death Eliminates Unwanted Cells". Molecular Biology of the Cell  (textbook) (5th ed.). Garland Science. p. 1115.ISBN 978-0-8153-4105-5

Apoptosis Interest Group (1999). "About apoptosis". Archived from the  original on 28 December 2006. Retrieved 2006-12-15.Webster.com  dictionary entry

Barichella M, Cereda E, Pezzoli G (October 2009). "Major nutritional issues in the management of Parkinson's disease". Mov. Disord. 24 (13): 1881–92.

Bronstein JM, Tagliati M, Alterman RL, et al. (October 2010). "Deep Brain Stimulation for Parkinson Disease: An Expert Consensus and Review of Key Issues". Arch Neurol 68 (2): 165–165.

 Bronstein JM, Tagliati M, Alterman RL, et al. (October 2010). "Deep Brain Stimulation for Parkinson Disease: An Expert Consensus and Review of Key Issues". Arch Neurol 68 (2): 165–165.

Brüne B (August 2003). "Nitric oxide: NO apoptosis or turning it ON?. Cell  Death Differ. 10 (8):  doi:10.1038/sj.cdd.4401261. PMID 12867993.

Brune, B. et al. 1999. "Nitric oxide (NO): an effector of apoptosis." Cell Death and Differentiation. 6:10 pg 969-975.

Bullani RR, Wehrli P, Viard-Leveugle I, Rimoldi D, Cerottini JC, Saurat JH, Tschopp J, French LE. Frequent downregulation of Fas (CD95) expression and function in melanoma. Melanoma Res. 2002;12:263–270. 

Burz C, Berindan-Neagoe I, Balacescu O, Irimie A. Apoptosis in cancer: key molecular signaling pathways and therapy targets. Acta Oncol. 2009;48:811–821. 

Butler LM, Hewett PJ, Butler WJ, Cowled PA. Down-regulation of Fas gene expression in colon cancer is not a result of allelic loss or gene rearrangement. Br J Cancer. 1998;77:1454–1459.[PMC free article

Ceravolo R, Frosini D, Rossi C, Bonuccelli U (December 2009). "Impulse control disorders in Parkinson's disease: definition, epidemiology, risk factors, neurobiology and management". Parkinsonism Relat. Disord. 15 Suppl 4: S111–5. doi:10.1016/S1353-8020(09)70847-8. PMID 20123548.

Ceravolo R, Frosini D, Rossi C, Bonuccelli U (December 2009). "Impulse control disorders in Parkinson's disease: definition, epidemiology, risk factors, neurobiology and management". Parkinsonism Relat. Disord. 15 Suppl 4: S111–5.

Chen G, Bhojani MS, Heaford AC, Chang DC, Laxman B, Thomas DG, Griffin LB, Yu J, Coppola JM, Giordano TJ, Lin L, Adams D, Orringer MB, et al. Phosphorylated FADD induces NF-kappaB, perturbs cell cycle, and is associated with poor outcome in lung adenocarcinomas.Proc Natl Acad Sci U S A. 2005;102:12507–12512.

Chiarugi A, Moskowitz  MA 2002. "PARP-1—a perpetrator of apoptotic cell death?".Science 297 (5579)

Chipuk JE, Green DR. How do BCL-2 proteins induce mitochondrial outer membrane permeabilization? Trends Cell Biol. 2008;18:157–164.

Chung, KA.; Lobb, B. M.; Nutt, J. G.; Horak, F. B. (October 2010). "Effects of    a central cholinesterase inhibitor on reducing falls in Parkinson disease". Neurology 75 (10): 1263–1269. doi:10.1212/WNL.0b013e3181f6128c.

Chung, KA.; Lobb, B. M.; Nutt, J. G.; Horak, F. B. (October 2010). "Effects of a central cholinesterase inhibitor on reducing falls in Parkinson disease". Neurology 75 (10): 1263–1269.

Cicchetti F, Drouin-Ouellet J, Gross RE (September 2009). "Environmental toxins and Parkinson's disease: what have we learned from pesticide-induced animal models?". Trends Pharmacol. Sci. 30 (9): 475–83.

Coffey RJ (March 2009). "Deep brain stimulation devices: a brief technical history and review". Artif Organs 33 (3): 208–20.

Collaborating Centre for Chronic Conditions, ed. (2006). "Symptomatic pharmacological therapy in Parkinson’s disease". Parkinson's Disease. London: Royal College of Physicians. pp. 59–100. ISBN 1-86016-283-5.

Dejean LM, Martinez-Caballero S, Manon S, Kinnally KW (February 2006). Regulation of the mitochondrial apoptosis-induced channel, MAC, by BCL-2 family proteins".Biochim. Biophys. Acta 1762 (2): 191–201. 

Dimond PF (2010-08-16), "No New Parkinson Disease Drug Expected  Anytime Soon", GEN news highlights (GEN-Genetic Engineering & Biotechnology News), retrieved 2010-10-25

Dixon L, Duncan D, Johnson P, et al. (2007). "Occupational therapy for patients with Parkinson's disease". In Deane, Katherine. Cochrane Database Syst Rev (3)

Dixon L, Duncan D, Johnson P, et al. (2007). "Occupational therapy for patients with Parkinson's disease". In Deane, Katherine. Cochrane Database Syst Rev (3): CD002813.

Donepezil (Aricept) Reduces Falls in People with Parkinson’s. Parkinson's Disease Foundation Science News. 11 November 2010.

Duan H, Chinnaiyan AM, Hudson PL, etal. 1996. ICE-LAP3, a novel                               mammalian homologue of the Caenorhabditis elegans cell death protein     Ced-3  is activated during Fas- and tumor necrosis factor-induced apoptosis. J. Biol. Chem. 271:1621–25 

Eberhard Y, Gronda M, Hurren R, Datti A, MacLean N, Ketela T, Moffat J, Wrana JL, Schimmer AD. Inhibition of SREBP1 sensitizes cells to death ligands. Oncotarget. 2011;2:186–196.

Egle A, Harris AW, Bath ML, O'Reilly L, Cory S. VavP-Bcl2 transgenic mice develop follicular lymphoma preceded by germinal center hyperplasia. Blood. 2004; 103:2276–2283. 

El Hindy N, Bachmann HS, Lambertz N, Adamzik M, Nuckel H, Worm K, Zhu Y, Sure U, Siffert W, Sandalcioglu IE. Association of the CC genotype of the regulatory BCL2 promoter polymorphism (-938C>A) with better 2-year survival in patients with glioblastoma multiforme.J Neurosurg. 2011;114:1631–1639. [PubMed]

Esposti MD. Bcl-2 antagonists and cancer: from the clinic, back to the bench. Cell Death Dis.2010;1:e37. [PMC free article] [PubMed]

Fahn S (2008). "The history of dopamine and levodopa in the treatment of Parkinson's disease". Mov. Disord. 23 Suppl 3: S497–508. doi:10.1002/mds.22028. PMID 18781671.

Fesik SW, Shi Y. (2001). "Controlling the caspases". Science 294 (5546): 1477–8.

Fox CM, Ramig LO, Ciucci MR, Sapir S, McFarland DH, Farley BG (November 2006). "The science and practice of LSVT/LOUD: neural plasticity-principled approach to treating individuals with Parkinson disease and other neurological disorders". Seminars in Speech and Language 27 (4): 283–99.

Fox CM, Ramig LO, Ciucci MR, Sapir S, McFarland DH, Farley BG (November 2006). "The science and practice of LSVT/LOUD: neural plasticity-principled approach to treating individuals with Parkinson disease and other neurological disorders". Seminars in Speech and Language 27 (4): 283–99.

Fox CM, Ramig LO, Ciucci MR, Sapir S, McFarland DH, Farley BG. The science and practice of LSVT/LOUD: neural plasticityprincipled approach to treating individuals with Parkinson’s disease disease and other neurological disorders. Semin Speech Lang 2006.

Fox CM, Ramig LO, Ciucci MR, Sapir S, McFarland DH, Farley BG. The science and practice of LSVT/LOUD: neural plasticityprincipled approach to treating individuals with Parkinson’s disease disease and other neurological disorders. Semin Speech Lang 2006;27:283-299.

Galluzzi L, Vitale I, Abrams JM, Alnemri ES, Baehrecke EH, Blagosklonny MV, Dawson TM, Dawson VL, El-Deiry WS, Fulda S, Gottlieb E, Green DR, Hengartner MO, et al. Molecular definitions of cell death subroutines: recommendations of the Nomenclature Committee on Cell Death 2012. Cell Death Differ. 2011 

Gascoyne RD, Adomat SA, Krajewski S, Krajewska M, Horsman DE, Tolcher AW, O'Reilly SE, Hoskins P, Coldman AJ, Reed JC, Connors JM. Prognostic significance of Bcl-2 protein expression and Bcl-2 gene rearrangement in diffuse aggressive non-Hodgkin's lymphoma.Blood. 1997;90:244–251.

Gavathiotis E, Walensky LD. Tracking BAX once its trigger is pulled. Cell Cycle. 2011;10:868–870. 

Gobe G, Rubin M, Williams G, Sawczuk I, Buttyan R. Apoptosis and expression of Bcl-2, Bcl-XL, and Bax in renal cell carcinomas. Cancer Invest. 2002;20:324–332. [PubMed]

Goldberg AA, Beach A, Davies GF, Harkness TA, Leblanc A, Titorenko VI. Lithocholic bile acid selectively kills neuroblastoma cells, while sparing normal neuronal cells. Oncotarget.2011;2:761–782. 

Goldenberg MM (October 2008). "Medical management of Parkinson's disease". P & T 33 (10): 590–606. PMC 2730785. PMID 19750042.

Goodwin VA, Richards SH, Taylor RS, Taylor AH, Campbell JL (April 2008).   "The effectiveness of exercise interventions for people with Parkinson's disease: a systematic review and meta-analysis". Movement Disorders 23 (5): 631–40.

Goodwin VA, Richards SH, Taylor RS, Taylor AH, Campbell JL (April 2008). "The effectiveness of exercise interventions for people with Parkinson's disease: a systematic review and meta-analysis". Movement Disorders 23 (5): 631–40.

Goodwin, V. A., Richards, S. H., Taylor, R. S., Taylor, A. H., Campbell, J. L. (2008). The effectiveness of exercise interventions for people with Parkinson’s disease: a systematic review and meta-analysis. Movement disorders, Vol. 23 (No. 5), 631-640.

Goodwin, V. A., Richards, S. H., Taylor, R. S., Taylor, A. H., Campbell, J. L. (2008). The effectiveness of exercise interventions for people with Parkinson’s disease: a systematic review and meta-analysis. Movement disorders, Vol. 23 (No. 5), 631-640.

Gratas C, Tohma Y, Barnas C, Taniere P, Hainaut P, Ohgaki H. Up-regulation of Fas (APO-1/CD95) ligand and down-regulation of Fas expression in human esophageal cancer. Cancer Res. 1998;58:2057–2062. [PubMed]

Green DR, Kroemer G. The pathophysiology of mitochondrial cell death. Science.2004;305:626–629.

Green, Douglas 2011. Means to an End: Apoptosis and other Cell Death  Mechanisms. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory         Press. ISBN 978-0-87969-888-1.

Guridi J, Lozano AM (November 1997). "A brief history of pallidotomy". Neurosurgery 41 (5): 1169–80; discussion 1180–3. doi:10.1097/00006123-199711000-00029. PMID 9361073.

Hahne M, Rimoldi D, Schroter M, Romero P, Schreier M, French LE, Schneider P, Bornand T, Fontana A, Lienard D, Cerottini J, Tschopp J. Melanoma cell expression of Fas(Apo-1/CD95) ligand: implications for tumor immune escape. Science. 1996;274:1363–1366.

Han H, Landreneau RJ, Santucci TS, Tung MY, Macherey RS, Shackney SE, Sturgis CD, Raab SS, Silverman JF. Prognostic value of immunohistochemical expressions of p53, HER-2/neu, and bcl-2 in stage I non-small-cell lung cancer. Hum Pathol. 2002;33:105–110.

Harvey BK, Wang Y, Hoffer BJ (2008). "Transgenic rodent models of  Parkinson's disease". Acta Neurochir. Suppl. 101: 89–92. doi:10.1007/978-3-211-78205-7_15. PMC 2613245. PMID 18642640.

Hasnain M, Vieweg WV, Baron MS, Beatty-Brooks M, Fernandez A, Pandurangi AK (July 2009). "Pharmacological management of psychosis in elderly patients with parkinsonism". Am. J. Med. 122 (7): 614–22. doi:10.1016/j.amjmed.2009.01.025. PMID 19559160.

Hasnain M, Vieweg WV, Baron MS, Beatty-Brooks M, Fernandez A, Pandurangi AK (July 2009). "Pharmacological management of psychosis in elderly patients with parkinsonism". Am. J. Med. 122 (7): 614–22.

Hornykiewicz O (2002). "L-DOPA: from a biologically inactive amino acid to a successful therapeutic agent". Amino Acids 23 (1–3): 65–70.

Jazirehi AR. Regulation of apoptosis-associated genes by histone deacetylase inhibitors: implications in cancer therapy. Anticancer Drugs. 2010;21:805–813. 

Johnstone RW, Frew AJ, Smyth MJ. The TRAIL apoptotic pathway in cancer onset, progression and therapy. Nat Rev Cancer. 2008;8:782–798.

Karam, Jose A. 2009. Apoptosis in Carcinogenesis and Chemotherapy. Netherlands: Springer. ISBN 978-1-4020-9597-9

Katzenschlager R, Evans A, Manson A, et al (2004). "Mucuna pruriens in  Parkinson's disease: a double blind clinical and pharmacological study". J. Neurol. Neurosurg. Psychiatr. 75 (12): 1672–7. doi:10.1136/jnnp.2003.028761. PMC 1738871. PMID 15548480.

Kaufmann T, Strasser A, Jost PJ. Fas death receptor signalling: roles of Bid and XIAP. Cell Death Differ. 2012;19:42–50. 

Kavathia N, Jain A, Walston J, Beamer BA, Fedarko NS. Serum markers of apoptosis decrease with age and cancer stage. Aging (Albany NY) 2009;1:652–663.

Kelly PN, Strasser A. The role of Bcl-2 and its pro-survival relatives in tumourigenesis and cancer therapy. Cell Death Differ. 2011;18:1414–1424. 

Kerr JF, Wyllie AH, Currie AR 1972. "Apoptosis: a basic biological            phenomenon with wide-ranging implications in tissue kinetics". Br. J.  Cancer

Knight RA, Melino G. Cell death in disease: from 2010 onwards. Cell Death Dis. 2011.

Koch G (2010). "rTMS effects on levodopa induced dyskinesias in Parkinson's disease patients: searching for effective cortical targets". Restor. Neurol. Neurosci. 28 (4): 561–8.

Ladha SS, Walker R, Shill HA (May 2005). "Case of neuroleptic malignant-like syndrome precipitated by abrupt fava bean discontinuance". Mov. Disord. 20 (5): 630–1. doi:10.1002/mds.20380. PMID 15719433.

Langston JW, Ballard P, Tetrud JW, Irwin I (February 1983). "Chronic Parkinsonism in humans due to a product of meperidine-analog synthesis". Science 219 (4587): 979–80.

Lanska DJ (2010). "Chapter 33: the history of movement disorders". Handb Clin Neurol 95: 501–46. doi:10.1016/S0072-9752(08)02133-7. PMID 19892136.

 Laurent M. Dejean, Sonia Martinez-Caballero, Kathleen W. Kinnally (2006). "Is MAC the knife that cuts cytochrome c from mitochondria during apoptosis?". Cell Death and Differentiation 13 (8)

Lavrik IN, Krammer  PH. Regulation of CD95/ Fas signalling at the DISC. Cell Death Differ.2012;19:36–41. 

Lee MS, Ernst E (January 2009). "Qigong for movement disorders: A systematic review". Mov. Disord. 24 (2): 301–3. doi:10.1002/mds.22275. PMID 18973253.

Lee MS, Lam P, Ernst E (December 2008). "Effectiveness of tai chi for Parkinson's disease: a critical review". Parkinsonism Relat. Disord. 14 (8): 589–94. doi:10.1016/j.parkreldis.2008.02.003. PMID 18374620.

Lee MS, Shin BC, Kong JC, Ernst E (August 2008). "Effectiveness of acupuncture for Parkinson's disease: a systematic review". Mov. Disord. 23 (11): 1505–15. doi:10.1002/mds.21993. PMID 18618661.

 Liu Y, Lang F, Xie X, Prabhu S, Xu J, Sampath D, Aldape K, Fuller G, Puduvalli VK. Efficacy of adenovirally expressed soluble TRAIL in human glioma organotypic slice culture and glioma xenografts. Cell Death Dis. 2011

Luthi AU, Martin SJ. The CASBAH: a searchable database of caspase substrates. Cell Death Differ. 2007. 

Mohan S, Bustamam A, Abdelwahab SI, Al-Zubairi AS, Aspollah M, Abdullah R, Elhassan MM, Ibrahim MY, Syam S: Typhonium flagelliforme induces apoptosis in CEMss cells via activation of caspase-9, Journal of ethnopharmacology 2010

Neznanov N, Komarov AP, Neznanova L, Stanhope-Baker P, Gudkov AV. Proteotoxic stress targeted therapy (PSTT): induction of protein misfolding enhances the antitumor effect of the proteasome inhibitor bortezomib. Oncotarget. 2011;2:209–221.

Obeso JA, Rodriguez-Oroz MC, Goetz CG, et al. (May 2010). "Missing pieces in the Parkinson's disease puzzle". Nat Med 16 (6): 653–61.

 Olsson M, Zhivotovsky B. Caspases and cancer. Cell Death Differ. 2011;18:1441–1449.[PMC free article] [PubMed] The National

O'Rourke MG, Ellem KA 2000. "John Kerr and apoptosis". Med. J. Aust. 173 (11–12): 616–7. PMID 11379508.

Parkinson's disease surgery neurology Channel. Retrieved on 2010-02-02Barichella M, Cereda E, Pezzoli G (October 2009). "Major nutritional issues in the management of Parkinson's disease". Mov. Disord. 24 (13): 1881–92.

Parkinson's disease surgery neurology Channel. Retrieved on 2010-02-02

Placzek WJ, Wei J, Kitada S, Zhai D, Reed JC, Pellecchia M. A survey of the anti-apoptotic Bcl-2 subfamily expression in cancer types provides a platform to predict the efficacy of Bcl-2 antagonists in cancer therapy. Cell Death Dis. 2010;1:e40. [PMC free article] 

Platz T, Rothwell JC (2010). "Brain stimulation and brain repair--rTMS: from animal experiment to clinical trials--what do we know?". Restor. Neurol. Neurosci. 28 (4): 387–98.

Raguthu L, Varanese S, Flancbaum L, Tayler E, Di Rocco A (October 2009). "Fava beans and Parkinson's disease: useful 'natural supplement' or useless risk?". Eur. J. Neurol. 16 (10): e171. doi:10.1111/j.1468-1331.2009.02766.x. PMID 19678834.

Rampino N, Yamamoto H, Ionov Y, Li Y, Sawai H, Reed JC, Perucho M. Somatic frameshift mutations in the BAX gene in colon cancers of the microsatellite mutator phenotype. Science.1997;275:967–969.

Samii A, Nutt JG, Ransom BR (May 2004). "Parkinson's disease". Lancet 363 (9423): 1783–93. doi:10.1016/S0140-6736(04)16305-8. PMID 15172778.

Samii A, Nutt JG, Ransom BR (May 2004). "Parkinson's disease". Lancet 363 (9423): 1783–93.

Shin MS, Kim HS, Lee SH, Lee JW, Song YH, Kim YS, Park WS, Kim SY, Lee SN, Park JY, Lee JH, Xiao W, Jo KH, et al. Alterations of Fas-pathway genes associated with nodal metastasis in non-small cell lung cancer. Oncogene. 2002;21:4129–4136. [PubMed]

Strand S, Hofmann WJ, Hug H, Muller M, Otto G, Strand D, Mariani SM, Stremmel W, Krammer PH, Galle PR. Lymphocyte apoptosis induced by CD95 (APO-1/Fas) ligand-expressing tumor cells--a mechanism of immune evasion? Nat Med. 1996;2:1361–1366.[PubMed]

Suchowersky O, Gronseth G, Perlmutter J, Reich S, Zesiewicz T, Weiner WJ (April 2006). "Practice Parameter: neuroprotective strategies and alternative therapies for Parkinson disease (an evidence-based review): report of the Quality Standards Subcommittee of the American Academy of Neurology". Neurology 66 (7): 976–82. doi:10.1212/01.wnl.0000206363.57955.1b. PMID 16606908.

Tadokoro D, Takahama S, Shimizu K, Hayashi S, Endo Y, Sawasaki T. Characterization of a caspase-3-substrate kinome using an N- and C-terminally tagged protein kinase library produced by a cell-free system. Cell Death Dis. 2010;1:e89. 

Tawfik K, Kimler BF, Davis MK, Fan F, Tawfik O. Prognostic significance of Bcl-2 in invasive mammary carcinomas: a comparative clinicopathologic study between ‘triple-negative’ and non-‘triple-negative’ tumors. Hum Pathol. 2011 

The National Collaborating Centre for Chronic Conditions, ed. (2006). "Surgery for Parkinson’s disease". Parkinson's Disease. London: Royal College of Physicians. pp. 101–111. ISBN 1-86016-283-5.

The National Collaborating Centre for Chronic Conditions, ed. (2006). "Other   key interventions". Parkinson's Disease. London: Royal College of Physicians. pp. 135–146. ISBN 1-86016-283-5.

The National Collaborating Centre for Chronic Conditions, ed. (2006). "Palliative care in Parkinson’s disease". Parkinson's Disease. London: Royal College of Physicians. pp. 147–151. ISBN 1-86016-283-5.

The National Collaborating Centre for Chronic Conditions, ed. (2006). "Symptomatic pharmacological therapy in Parkinson’s disease". Parkinson's Disease. London: Royal College of Physicians. pp. 

The National Collaborating Centre for Chronic Conditions, ed. (2006). "Non-motor features of Parkinson’s disease". Parkinson's Disease. London: Royal College of Physicians. pp. 113–133. ISBN 1-86016-283-5.

The National Collaborating Centre for Chronic Conditions, ed. (2006). "Surgery for Parkinson’s disease". Parkinson's Disease. London: Royal College of Physicians. pp. 101–111. ISBN 1-86016-283-5.

The National Collaborating Centre for Chronic Conditions, ed. (2006). "Other key interventions". Parkinson's Disease. London: Royal College of Physicians. pp. 135–146. ISBN 1-86016-283-5.

The National Collaborating Centre for Chronic Conditions, ed. (2006). "Palliative care in Parkinson’s disease". Parkinson's Disease. London: Royal College of Physicians. pp. 147–151.

Ueno T, Toi M, Tominaga T. Circulating soluble Fas concentration in breast cancer patients.Clin Cancer Res. 1999;5:3529–3533. [PubMed]

Viard-Leveugle I, Veyrenc S, French LE, Brambilla C, Brambilla E. Frequent loss of Fas expression and function in human lung tumours with overexpression of FasL in small cell lung carcinoma. J Pathol. 2003;201:268–277. 

Wickman G, Julian L, Olson MF. How apoptotic cells aid in the removal of their own cold dead bodies . Cell Death Differ. 2012;19: 735–742. 

Yin C, Knudson CM, Korsmeyer SJ, Van Dyke T. Bax suppresses tumorigenesis and stimulates apoptosis in vivo. Nature. 1997;385:637–640.

Yivgi-Ohana N, Eifer M, Addadi Y, Neeman M, Gross A. Utilizing mitochondrial events as biomarkers for imaging apoptosis. Cell Death Dis. 2011;2:e166.

Zinkel SS, Hurov KE, Ong C, Abtahi FM, Gross A, Korsmeyer SJ. A role for proapoptotic BID in the DNA-damage response. Cell. 2005;122:579–591.