Introduction And Outline Of The Paraganglion System Biology Essay

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Paraganglia are small bodies of chromophil cell clusters associated with the ganglia of the autonomic nervous system. The paraganglion system consists of the adrenal medulla, the largest paraganglion in the human body, the orthosympathetic paraganglia and the parasympathetic paraganglia. The orthosympathetic paraganglia are localized in the prevertebral and paravertebral orthosympathetic paraganglia, the organ of Zuckerkandl , along the hypogastric plexus and along the urinary bladder.

Fig.1 The adrenal medulla and extra-adrenal orthosympathetic paraganglia.

The parasympathetic paraganglia consist of the intravagal bodies and the branchiomeric paraganglia in the mediastinum and head and neck region, most notably located in the carotid bifurcation, the jugular foramen and on the promontory of the middle ear.

Fig.2 The parasympathetic branchiomeric paraganglia

Paraganglia consist of a parenchymal component and a stromal component. The parenchymal component is of neuroectodermal origin and gives rise to the type 1 or chief-cells within the paraganglia. During embrygenesis it is thought to migrate along nerves or vasculature from the neural crest to the locations along the cranial nerves, sympathetic trunk and greater vessels

mentioned above. The stromal component is of mesenchymal origin en contains the type 2 or sustentacular cells, as well as other stromal components such as blood vessels. Type 1 and 2 cells form a specific configuration known as the "zellballen": the chief cells or type 1 cells form small clusters surrounded by type 2 cells and other stromal components. The exact function of the paraganglion system is unknown, except for the carotid and aortic bodies, which function as chemoreceptors sensitive to changes in arterial pH and oxygen tension and play a role in the control of cardiovascular and respiratory centers.


type 1 cells

type 2 cells

Figure 1. microscopy of hematoxylin and eosin (H-E) stained paraganglion tissue showing the type 1 and 2 cells in the classic Zellballen configuration.

Neoplasia of the paraganglion system

The nomenclature for the neoplasia arising from the paraganglion system is unequivocal and has changed over time. The terms 'glomus tumors', 'chemodectomas', 'paragangliomas' and 'phaeochromocytomas' have all been used interchangeably. The current classification according to the World Health Organization (WHO) designates tumors arising in the parasympathetic paraganglia in the head and neck region as 'paragangliomas', accompanied by the site of origin, i.e 'carotid body paraganglioma'. The term 'phaeochromocytoma' is reserved for tumors arising in the adrenal medulla, and 'extra-adrenal paraganglioma' for tumors arising in orthosympathetic paraganglia elsewhere in the retroperitonal space, the abdomen, or in the thorax{Tischler, 2008 191 /id}. Confusingly, the term phaeochromocytoma is sometimes used for all orthosympathetic paraganglion tumors. Paragangliomas are usually slow growing, highly vascular tumors. The typical architecture of normal paraganglion tissue, the 'zellballen' configuration consisting of type 1 and type 2 cells, is usually maintained in the tumor, although in pheochromocytomas, it may be less prominent {Strong, 2008 133 /id;Lack, 1979 71 /id}{Tischler, 2008 191 /id}. It has been demonstrated that the tumorigenic component is formed by the type 1 or chief cells, and that the type 2 stromal cells show expansion under the influence of the type 1 cells{Douwes Dekker, 2004 54 /id}.

Paragangliomas of the head and neck

Paragangliomas of the head and neck are rare tumors, comprising approximately 0.6% of all head and neck neoplasms{Batsakis, 1979 189 /id}. The incidence is estimated to be between 1:1.000.000 and 1:100.000{Lack, 1979 71 /id;Oosterwijk, 1996 81 /id;van der Kleij-Corssmit EP, 2008 166 /id}. These figures are based on pooled data from Dutch pathology laboratories and surgical patients. Due to the benign natural course of the disease, paragangliomas will not be operated upon in sizable proportion of patients and it is therefore likely that these figures represent an underestimation of the actual incidence{Baysal, 2002 13 /id;van der Kleij-Corssmit EP, 2008 166 /id}. Necroscopy rates for carotid body paragangliomas of 1:3.860 to 1:13.4000 also point towards a higher incidence and suggest that a significant number of paragangliomas are not surgically removed{Baysal, 2002 13 /id;van der Kleij-Corssmit EP, 2008 166 /id}. The incidence of paragangliomas seems to be influenced by environmental factors that facilitate paraganglioma formation, such as high altitude (see alo paragraph…) or genetic factors such as the regional clustering of paraganglioma patients due to a common hereditary genetic trait, as can be seen in the Netherlands (see also the paragraph 'genetics of paraganglioma', and chapters two and three). Head and neck paragangliomas most frequently arise in the carotid bifurcation as carotid body tumors (in approximately 80%). Other common locations within the head and neck region are along the jugular bulb or tympanic nerve (in approximately 17.5%), or along the vagal nerve (in approximately 4.5%). Symptoms of head and neck pararagangliomas vary with the tumor localization. Most tumors are characterized by slow expansive growth, resulting in a non-painful, palpable neck mass or pharyngeal bulging. Cranial nerve dysfunction may occur, especially of the facial, glossopharyngeal, vagal, accessory and hypoglossal nerves because of their close anatomical relations with the jugulotympanic- , vagal- and carotid paraganglia. In case of tympanic or jugulotympanic tumors, there may be conductive hearing loss and tinnitus, which is pulsatile in typical cases. In case of functional paragangliomas, patients can present which symptoms and signs of catecholamine excess (see below).

The diagnosis of head and neck paragangliomas is based on the patient- and family history, clinical investigation of the ears, the pharynx and the neck, biochemical screening for catecholamine excess (see below), and radiology. In the head and neck region, both high resolution computed tomography (CT) and magnetic resonance imaging (MRI) are sensitive modalities to detect tumor masses. Whereas they lack specificity for paragangliomas, they are useful in assessing the extent and anatomical relations of the tumor, and detect multiple lesions within the head and neck region if present. Some argue that the preferred imaging modality is MRI, because contrast used in CT imaging might provoke catecholamine release in patients that are not pre-treated with alpha- or beta-blockers, however recent data contradict this finding {Raisanen, 1984 172 /id;Bessell-Browne, 2007 171 /id;Baid, 2009 174 /id}. Functional imaging techniques like 131I- metaiodobenzylguanidine (MIBG) scintigraphy, 18F-fluorodopamine or 18F -fluorodihydroxyphenylalanine positron emission tomography (18F-FDA PET and 18F-FDOPA PET) have a higher specificity because they detect abnormal isotope uptake by norepinephrine transporters in paraganglioma tissue{Chen, 2010 175 /id;Kantorovich, 2010 165 /id}. They are useful when in doubt of the diagnosis and in whole-body screening for functional paragangliomas and phaeochromocytomas. The disadvantage is the lack of anatomical detail in the images, and a reduced sensitivity of MIBG and 18F-FDA PET has been described in extra-adrenal and malignant paraganglioma{Chen, 2010 175 /id;Kantorovich, 2010 165 /id;Petri, 2009 167 /id}. Last, 18F-fluorodeoxyglucose (FDG) PET is efficient in whole-body screening for metabolically active tissue. As such, it is not very specific for paragangliomas or phaeochromocytomas, but it is useful in screening for multiple tumors and has been shown to be a superior tool in the detection of paraganglioma metastases{Chen, 2010 175 /id;Kantorovich, 2010 165 /id}. Angiography can identify paragangliomas as highly vascular lesions, and assess the vascular anatomy and main contributing blood vessels. It is especially useful if surgery is considered and pre-operative elimination of the blood supply to the tumor is warranted. Definitive confirmation of the diagnosis is obtained by histopathology and the identification of the pathognomonic 'Zellballen' configuration within the tumor tissue. However, because of the high vascularity of the tumors and the risk of profuse bleeding upon biopsy, tissue samples for histopathology are rarely available prior to the surgical resection of the tumor.

In general, there are three strategies for the management of head and neck paragangliomas. The first is surgical resection of the paraganglioma. The obvious benefit of this strategy is the complete removal of the tumor. However, due to the high vascularity of paragangliomas and their close anatomical relationships with the carotid artery, the jugular vein, multiple cranial nerves, and/or the skull base, there is a definite risk of surgical complications, especially in larger tumors and those invading the skull base{van der Kleij-Corssmit EP, 2008 166 /id;Papaspyrou, 2009 169 /id}. The second option, radiotherapy, has a much reduced risk of bleeding and cranial nerve injury{Foote, 2002 170 /id}. Its aim is local control of tumor growth, and the disadvantages therefore are the persistence of the tumor, the possible long-term effects of irradiation and the greater surgical difficulty and risk if resection proves to be necessary at a later stage. The third option consists of a policy of watchful waiting, or 'wait and scan'. No intervention is performed, and tumor growth is monitored regularly with repeated CT or MR imaging. Surgery or radiotherapy is undertaken only if there is evidence of tumor growth or impending complications. The disadvantage of this strategy is the persistence of the tumor and its potential to grow, however, most head and neck paragangliomas are characterized by an indolent growth pattern, a substantial number of head and neck paragangliomas will not become symptomatic, and the effects of cranial nerve palsy are often better mediated if the paresis is slowly progressive due to tumor growth as opposed to sudden paralysis due to surgical injury{Hensen, 2009 111 /id}. Not surprisingly, the optimal management strategy for head and neck paragangliomas is subject of much debate in the literature{Huy, 2009 173 /id;Gjuric, 2009 160 /id;Mendenhall, 2010 164 /id;Jansen, 2000 87 /id;van der Mey, 1992 70 /id}. In the Netherlands, the choice of treatment modality and timing are tailored to the individual patient and depend on tumor location, tumor size, mutlifocality, catecholamine excess (see below), and the causative gene mutation{van der Kleij-Corssmit EP, 2008 166 /id}. Watchful waiting is often deemed appropriate for patients with multiple, asymptomatic, or very large skull base paragangliomas, those of advanced age, and in case of gene mutations characterized by a mild disease phenotype{van der Kleij-Corssmit EP, 2008 166 /id}.

Phaeochromocytomas and extra-adrenal paragangliomas

Phaeochromocytomas and extra-adrenal paragangliomas are tumors arising in the adrenal medulla and sympathetic paraganglia respectively, and are commonly described as sympathetic paragangliomas. Phaeochromocytomas and extra-adrenal paragangliomas are rare tumors, with an estimated incidence between 0,2:100.000 and 2:100.000 {Pacak, 2001 179 /id;Harding, 2005 180 /id}. In about 80%, they form in the adrenal medulla, approximately 20% occur elsewhere from the neck down to the pelvic floor in any of the sympathetic paraganglia, most frequently in the abdomen, the pelvis and less frequently in the thorax{Petri, 2009 167 /id}. Extra-adrenal sympathetic paragangliomas show a greater degree of malignancy than either phaeochromocytomas or head and neck paragangliomas {Benn, 2006 78 /id}. The symptoms of phaeochromocytomas are usually caused by the excretion of catecholamines or their metabolites by the tumor, and include hypertension (in about 60%), which may be fluctuating or sustained, paroxysmal palpitations, headache, agitation, excess sweating and pallor. Diagnosis of pahaeochromocytomas and extra-adrenal paragangliomas is based on biochemical screening for catecholamine excess and radiology. Both abdominal CT and MRI are sensitive modalities for the detection of abdominal masses. As described above, some authors prefer MRI because no iodine containing contrast is needed. The mainstay of phaeochromocytoma therapy is surgical resection. The preferred technique is adrenalectomy via a laparoscopic or retroperitoneoscopic approach, as this minimizes risk and surgical morbidity{Walz, 2006 184 /id;Brunt, 2002 183 /id;Brunt, 2006 182 /id}. In case of bilateral disease or a genetic risk of developing bilateral disease, bilateral cortical sparing adrenalectomies may be considered to preserve adrenal glococortocoid production{Brunt, 2006 182 /id}. Open laparotomy is sometimes necessary, especially in case of large tumors (diameter of 6 cm or more){Brunt, 2006 182 /id}. Because of the risk of catecholamine excess during, and catecholamine depletion after operation, peri-operative treatment with α- and β- ardrenoreceptor antagonists, calcium channel blockers and/or catecholamine synthesis-inhibitors are mandatory (see paragraph functional paragangliomas).

Functional paragangliomas

Neoplasia of the paraganglion system may excrete vasoactive catecholamines like dopamine, adrenalin and noradrenalin and/or their metabolites. This is a well-known feature of adrenal phaeochromocytomas, but rare in head and neck paragangliomas, of which only 1-5% is estimated to produce excess catecholamines{Erickson, 2001 158 /id} (Zak&Lawson 1982). The majority of functional paragangliomas produce noradrenalin, only very few secrete dopamine or adrenalin{Colen, 2009 147 /id;VAN DER Horst-Schrivers AN, 2010 144 /id}. The diagnosis of catecholamine excess is based on 24-hours urinary metanephrine and normetanephrine measurements, because of its high sensitivity and specificity {van der Kleij-Corssmit EP, 2008 166 /id}. This test should be performed in case of phaeochromocytomas, extra-adrenal paragangliomas, if a patient's signs or symptoms indicate a functional paraganglioma, or if a genetic risk exists for the development of paragangliomma-phaeochromocytoma syndrome (see below){Petri, 2009 167 /id;Chen, 2010 175 /id;van der Kleij-Corssmit EP, 2008 166 /id}. If catecholamine secretion is present in head and neck paragangliomas, it can cause the same symptoms as it does in pheaochromocytomas (hypertension, palpitations, headache, agitation, excess sweating and palor). In some cases, the catecholamine secretion causes serious hyperglycaemia or electrolyte disturbances. Prolonged exposure to high levels of catecholamines can cause cardiovascular complications such as cardiac hypertrophy, myocardial infarction, heart failure, and phaeochromocytomas are in rare cases associated with shock, multiple organ failure, and sudden death by stroke or cardiac arrest{Lenders, 2005 188 /id} {Galetta, 2010 168 /id;Petri, 2009 167 /id}. Because of these potentially life-threatening conditions, surgical excision is the treatment of choice in functional paragangliomas and phaeochromocytomas{Chen, 2010 175 /id;van der Kleij-Corssmit EP, 2008 166 /id}. Stringent intra-operative monitoring and peri-operative measures consisting of treatment with α- and β- ardrenoreceptor antagonists, calcium channel blockers and/or catecholamine synthesis-inhibitors, are mandatory to counter critical hypertensive crises and compensatory hypotensive episodes due to manipulation and removal of the tumor{Petri, 2009 167 /id;Brunt, 2006 182 /id}. Because of the clinical consequences of catecholamine secreting tumors, some authors propose to classify paragangliomas not according to their anatomical localization but according to whether or not they show functional activity{Neumann, 2009 129 /id}.


Most paragangliomas are benign tumors, i.e. they do not metastasize and are characterized by an expansive rather than an invasive growth pattern. However, some tumors, especially within the petrous bone, show erosion of the surrounding bone, some tumors show microvascular invasion, and some tumors do metastasize. As of yet, no definite histopathologic criteria for malignant paraganglioma have been established{Tischler, 2008 191 /id}. Even malignant paragangliomas and their metastases usually demonstrate the well differentiated growth pattern of normal paraganglion tissue{Strong, 2008 133 /id;Lack, 1979 71 /id}. Factors such as a higher mitotic rate, tumor cell spindling, altered nuclear morphology, aberrant DNA-ploidy, necrosis, and capsular or microvascular invasion are reported to be more prevalent in malignant cases, but are all also found in benign paragangliomas{Strong, 2008 133 /id;Lack, 1979 71 /id}{Tischler, 2008 191 /id}. Immunohistochemical markers such as Ki-67, Cyclin-D1, p53, p21, p27, BCL-2 and MDM-2 have been shown to be of little use in predicting malignancy in paraganglioma-phaeochromocytomas {Strong, 2008 133 /id}{Tischler, 2008 191 /id}. Malignant paraganglioma is therefore defined as metastatic paraganglioma in non-neuroendocrine tissue. The prevalence of metastatic paraganglioma varies in different patient series, from 0-10%. In case of matastatic head and neck paragangliomas, metastases are most frequently found in cervical lymph nodes (69%). Distant metastases are identified in 31% of malignant HNPGL, and the distant predilection sites include bone, lung and liver{Lee, 2002 137 /id}. Several studies have assessed clinical factors that may predict malignancy in paraganglioma patients. Features such as young age at diagnosis, pain as an accompanying symptom, rapidly enlarging tumor mass, large tumor size, and mediastinal and extra-adrenal abdominal tumor localization all seem to be associated with an increased risk of malignancy, but none of these features are proof of malignancy in themselves {Strong, 2008 133 /id;Ayala-Ramirez, 2010 135 /id;Chapman, 2010 146 /id}. Functional tumors that secrete catecholamines may be malignant or benign in nature. There is some debate as to whether dopamine secretion is indicative of extra-adrenal tumor localization and malignancy, but recent studies show that dopamine secretion is not uncommon in HNPGL (19-23%), and is not related to metastatic disease or outcome{Eisenhofer, 2005 151 /id;John, 1999 134 /id;van, 2010 148 /id;VAN DER Horst-Schrivers AN, 2010 144 /id}. The risk of metastatic disease is however correlated with the gene defect causing the paraganglioma (see paragraph genetics of head and neck paragangliomas).

The management of metastatic disease is challenging and .

More aggressive strategies are aimed at eradication and/or control of tumor growth both at the primary and metastatic site and include therapeutic embolization, surgical excision, radiotherapy, systemic chemotherapy (with cyclophosphamide, vincristine and dacarbazine), or combinations thereof{Moskovic, 2010 132 /id}{Lee, 2002 137 /id}.{Plouin, 2010 138 /id}. In case of symptomatic metastases that are too widespread for surgery or embolization, pharmacological blocking of catecholamine secretion, palliative conventional radiotherapy, metabolic radiotherapy with 131I-MIBG47, and/or systemic chemotherapy may be considered{Plouin, 2010 138 /id}{Lee, 2002 137 /id}{Moskovic, 2010 132 /id}. In stable metastatic disease, a policy of watchful waiting seems a viable option, as it is for stable primary disease{Plouin, 2010 138 /id}{Moskovic, 2010 132 /id}. There is some evidence that survival is improved by surgery in resectable metastatic disease, but due to the rarity of malignant paragangliomas and the number of different strategies and regimens that have been used over time, data on the outcome of these strategies are largely retrospective, not fully comparable, and often biased, and the lack of controlled prospective trials hampers the recommendation of specific therapeutic modalities{Moskovic, 2010 132 /id}{Plouin, 2010 138 /id}{Lee, 2002 137 /id}.

Genetics of head and neck paraganglioma

The knowledge of paraganglioma and pheochromocytoma genetics was, until the year 2000, limited to mutations in the VHL, RET and NF1 genes, causing Von Hippel-Lindau (VHL) syndrome, multiple endocrine neoplasia type 2a and 2b (MEN2a or MEN2b), or neurofibromatosis type 1 (NF1) respectively(Tischler, 2008). Whereas phaeochromocytomas are known to be part of the tumor spectrum of these syndromes, head and neck paragangliomas caused by VHL, RET or NF1 mutations are extremely rare. As this thesis focuses on head and neck paragangliomas, VHL, RET or NF1 mutations and their associated syndromes are not discussed in detail.

In 2000, Baysal et al. in collaboration with the Paraganglioma research Group Leiden, discovered that mutations in the SDHD gene, located on the long arm of chromosome 11 (11q23), cause hereditary head and neck paraganglioma syndrome type 1 (PGL1){Baysal, 2000 22 /id}. This break-through discovery of SDHD, a nuclear gene encoding an anchoring subunit of the mitochondrial succinate dehydrogenase complex (SDH), was soon followed by the identification of defects in other SDH subunits causing hereditary paraganglioma syndrome. In 2000, SDHC, located on chromosome 1 (1q23), encoding a another SDH anchoring subunit, was found to be the causative gene in paraganglioma syndrome PGL3{Niemann, 2000 31 /id}. In 2001, SDHB, located on chromosome 1 (1p35-36.1), encoding a catalytic iron-sulfur SDH subunit, was linked to paraganglioma syndrome PGL4{Astuti, 2001 25 /id}. Recently, new SDH-related genes have been associated with hereditary paragangliomas. SDHAF2 (formerly known as SDH5), located on chromosome 11 (11q13), encodes a SDH co-factor related to the function of the SDHA subunit, and is associated with head and neck paraganglioma syndrome PGL2{Hao, 2009 110 /id}. SDHA (located on 5p15), encoding the flavoprotein subunit of SDH, has itself now been associated with paragangliomas{Burnichon, 2010 185 /id}.

All SDH genes act as tumor suppressor genes. In paraganglioma patients carrying a germ-line mutation in a SDH gene, loss of heterozygosity (LOH), i.e. the loss of the normal allele, is required for tumorigenesis. Loss of the normal allele in conjunction with a germ-line mutation in a SDH gene results in the loss of the protein subunit, which destabilizes the SDH complex and disrupts its enzymatic activity{Douwes Dekker, 2003 127 /id}. Despite the fact that SDH proteins are all components of the same protein complex, mutations in different SDH genes lead to distinct clinical phenotypes. Mutations in SDHB, SDHC and SDHD, but not in SDHA or SDHAF2 are associated with the development of phaeochromocytomas. Mutations in SDHAF2 and SDHD show a high penetrance (75-100%) and frequently cause multiple head and neck paragangliomas (in 70-91%), whereas SDHB mutations show a lower penetrance (25-40%) and are more closely related to extra-adrenal paragangliomas, phaeochromocytomas and malignancy (see chapters 3 and 5){Kunst, 2011 159 /id;Neumann, 2004 77 /id;Benn, 2006 78 /id;Hes, 2010 120 /id;Ricketts, 2010 114 /id;Solis, 2009 116 /id;Schiavi, 2010 130 /id}. Mutations in SDHC are rare, and primarily associated with head and neck paragangliomas, although extra-adrenal paragangliomas, phaeochromocytomas and malignancy have been reported in SDHC-linked cases. {Schiavi, 2005 2 /id;Peczkowska, 2008 98 /id;Niemann, 2000 31 /id}. SDHA mutations have long not been associated with paragangliomas, but with severe disease phenotypes consisting of spasticity, cardiomyopathy, and neurodegeneration, until Burnichon et al. identified a mutation in SDHA in a single patient with an extra-adrenal paraganglioma{Burnichon, 2010 185 /id} (Horvath 2006) (Pagnamenta 2006)(Levitas 2010).

Currently, pathogenic gene mutations can be identified in approximately 32% of paraganglioma and pheochromocytoma patients{Mannelli, 2009 126 /id}. Mutations in VHL, RET and NF1 account for approximately 17% of the cases, but are very rarely associated with head and neck or extra-adrenal paragangliomas{Eisenhofer, 2011 149 /id}. Mutations in SDHB, SDHC, SDHD and SDHAF2 account for the remaining 15% (Mannelli, 2009). However, considerable differences in the relative mutation frequencies have been reported in different patient cohorts and different parts of the world. In the Netherlands, mutations in SDH genes, especially SDHD, seem to play a more prominent role than elsewhere, most likely due to the occurrence of Dutch founder mutations in SDHD: SDHD.D92Y, SDHD.L139P and SDHD.L95P (see also chapters 3 and 4){Dannenberg, 2002 107 /id;Taschner, 2001 17 /id}. It is likely that the high prevalence of founder mutations causes a higher prevalence of paragangliomas in the Netherlands (see also chapter 3){Baysal, 2008 186 /id}. For a more detailed description of the SDH genes and their associated tumor syndromes the reader is referred to chapters 2 and 3.



protein function



paraganglioma predilection site



neurofibromin, RAS pathway regulator

autosomal dominant

neurofibromatosis type 1 (NF1)




receptor tyrosine kinase

autosomal dominant

multiple endocrine neoplasia (MEN 2a/2b)




E3 ubiquitin ligase subunit

autosomal dominant

Von Hippel-Lindau syndrome (VHL)




flavoprotein catalytic subunit of SDH

autosomal recessive

Leigh syndrome, myopathy, encephalopathy




assembly factor of SDH

autosomal dominant, parent-of -origin dependent

paraganlioma syndrome (PGL2)

head and neck



iron-sulphur catalytic subunit of SDH

autosomal dominant

paraganlioma syndrome (PGL4)

extra-adrenal, adrenal, thoracic



anchoring subunit of SDH

autosomal dominant

paraganlioma syndrome (PGL3)

head and neck



anchoring subunit of SDH

autosomal dominant, parent-of -origin dependent

paraganlioma syndrome (PGL1)

head and neck

Table 1. Summary of paraganglioma-phaeochromocytoma genes and the associated syndromes

Inheritance of head and neck paraganglioma syndromes

The inheritance of paraganglioma syndrome differs significantly dependent on the gene involved. While SDHB- and SDHC-linked paraganglioma families show normal autosomal dominant inheritance, SDHD and SDHAF2 linked families show an exclusively paternal transmission of tumor susceptibility {van der Mey, 1989 69 /id;Hao} (Struycken 1997:Glomus tumors and genomic imprinting). Although mutations in SDHD and SDHAF2 can be inherited both via the maternal and paternal lines, tumor formation following maternal transmission of a mutation is extremely rare {Pigny, 2008 90 /id;van der Mey, 1989 69 /id}.

The failure of maternally transmitted mutations to initiate tumorigenesis initially suggested that only the paternal alleles of SDHD were expressed, a phenomenon consistent with maternal imprinting of the SDHD gene {van der Mey, 1989 69 /id}. However, it was established that the gene does not show mono-allelic expression in non-paraganglioma tissue {Baysal, 2000 19 /id;Hensen, 2004 4 /id}. The concept of maternal imprinting of SDHD also does not explain the frequent loss of heterozygosity at chromosome 11 that has been observed in SDHD-linked paragangliomas, because loss of the wild type allele would not constitute a predisposition to tumorigenesis if the wildtype allele was already inactivated by an imprint It is known that the entire maternal copy of chromosome 11 is lost in many paragangliomas {Hensen, 2004 4 /id;Dannenberg, 2001 50 /id;Yamashita, 2009 187 /id}. Furthermore, the actual blocking of transcription of the SDHD gene by methylation of the 11q23 region has never been reported.

As the exclusive paternal transmission of paragangliomas is also present in families linked to SDHAF2 (like SDHD located on the long arm of chromosome 11), while it is absent in SDHB- and SDHC-related tumors (both genes located on chromosome 1), the location of SDHAF2 and SDHD on chromosome 11 seems to be a decisive factor in the inheritance of SDHAF2 and SDHD linked paraganglioma syndrome. Whereas SDHD and SDHAF2 themselves do not seem to be imprinted, the main cluster of imprinted genes in the human genome is located on the same chromosome, at 11p15.5. This suggests a model in which a maternally expressed, paternally imprinted gene is an essential initiator or modifier of tumor development in these syndromes (see also chapter 7){Hensen, 2004 4 /id;Pigny, 2008 90 /id}. Indeed, the only report to date that has claimed to show the maternal transmission of tumor susceptibly together with an SDHD mutation showed that the patient had also acquired an altered methylation profile and therefore probably an altered imprinted status of H19, a known paternally imprinted tumor suppressor gene on 11p15 {Pigny, 2008 5814 /id;Yoshimizu, 2008 5789 /id}.In addition, it is known that VHL-related phaeochromocytomas {Margetts, 2005 5762 /id;Mircescu, 2001 5785 /id} also show loss of the maternal copy of the chromosome 11p15.5 region specifically, indicating that this model may have wider importance.

High Altitude Paraganglioma

Long before the identification of any of the genes now known to play a role in paraganglioma, it was recognized that living at high altitude can have a profound influence on the development of carotid body hyperplasia and carotid body tumors {Arias-Stella, 1969 5876 /id;Edwards, 1971 696 /id;Arias-Stella, 1973 5910 /id}. A number of mammalian species are known to develop pronounced hyperplasia or tumors with a prevalence of up to 10% in humans and up to 40% in bovines {Arias-Stella, 1976 5877 /id;Saldana, 1973 47 /id}, in contrast to an estimated low altitude prevalence of head and neck paraganglioma of 1 in 500,000 or less.

This increased prevalence and the central role of the carotid body in oxygen sensing suggested a role for oxygen sensing in the tumorigenesis of paragangliomas. The identification of succinate dehydrogenase and subsequent molecular studies has affirmed this link. A number of studies have linked the central mediator of cellular hypoxia, HIF-1, to defects in succinate dehydrogenase {King, 2006 5857 /id}. These studies postulate that a so-called 'pseudo-hypoxia' results from the inhibition of succinate dehydrogenase, leading to the accumulation of succinate, resulting in the activation of HIF-1 through the inhibition of prolyl hydroxylase-mediated degradation {Selak, 2005 5856 /id;Koivunen, 2007 5952 /id}. The HIF-1 transcription factor complex {Semenza, 2006 5917 /id} initiates the transcription of a range of genes that mediate an adaptive response to reduced oxygen. How the activation of the HIF-1 protein may lead to the initiation of tumorigenesis in the carotid body and the exact relation of physiological hypoxia to molecular 'pseudo-hypoxia' awaits further investigation. Despite this suggestive link, the possible role of succinate dehydrogenase mutations in high altitude paraganglioma cases has received little attention and the first genetic analysis failed to identify any mutations {Jech, 2006 5880 /id}. Recently, Cerecer-Gil et al. {Cerecer-Gil, 2010 6162 /id} identified a family with two SDHB-linked cases of high altitude paraganglioma, residing at elevations of up to 2200m. These are the first cases to link high altitude paraganglioma to mutations of the succinate dehydrogenase genes. While the occurrence of paraganglioma in this family could be purely coincidental to their place of residence, two factors indicated that elevation may be playing a role in the expression off these tumors. One of the patients showed a remarkably aggressive recurrent tumor, which achieved a volume almost equivalent to the original tumor within two months of excision. This behavior is in sharp contrast with the indolent growth pattern normally seen in head and neck paragangliomas, with a mean doubling rate of 4.2 years {Jansen, 2000 87 /id}. In addition, both patients developed head and neck tumors, while abdominal tumors occur much more frequently in SDHB mutation carriers. The identification of SDHB mutations in high altitude paraganglioma may serve to renew interest in this fascinating but underappreciated field of paraganglioma research, and refocus attention on the role of oxygen levels in the initiation and development of these tumors


Tumor biology of head and neck paragangliomas

Molecular biology of paragangliomas

Succinate dehydrogenase is an enzyme of the mitochondrial tricarboxylic acid cycle, and also plays an important role as the complex II component of the electron transport chain, contributing to the generation of ATP by oxidative phosphorylation. These combined roles place SDH at the center of two of the essential energy producing processes of the cell. SDHA is a flavoprotein, and SDHB, an iron-sulfur protein, and together they form the main catalytic domain, while SDHC and SDHD are the membrane-anchoring subunits of SDH and play a role in passing electrons through the electron transport chain.

Outline of the thesis

The aim of this thesis is to gain insight in the genetics, inheritance and tumor biology of paragangliomas and the clinical consequences for paraganglioma patients, with a focus on hereditary paraganglioma syndrome in the Netherlands. Chapter one consists of a general introduction into the current insights in the clinical characteristics, the genetics and tumor biology of paragangliomas and phaeochromocytomas.

In chapter two, the mutation frequency in succinate dehydrogenase genes in the Netherlands is analyzed, with the aim of uncovering the relative role of each of these genes in the Dutch paraganglioma population.

In chapter three, the clinical characteristics and the mutation frequency are evaluated of Dutch head and neck paraganglioma patients treated at the Leiden University Medical Center (LUMC). It describes the peculiar genetic make-up of the Dutch head and neck paraganglioma population and the consequences for the clinical picture of paraganglioma syndrome in the Netherlands.

In chapter four, the phenotype of the SDHD.D92Y Dutch founder mutation is studied in a large multigenerational paraganglioma kindred, with a focus on penetrance. As is shown in chapters two and three, the D92Y (or p.Asp92Tyr) mutation in SDHD is one of the major causes of paraganglioma in the Netherlands.

In chapter five, the unusual parent-of-origin dependent inheritance that is observed in all SDHD-linked paraganglioma kindreds, including the family described in chapter four, is further investigated. A hypothesis is put forward that explains the exclusive paternal transmission of pararangliomas in SDHD-linked families in the absence of evidence for maternal imprinting of the SDHD gene itself.

In chapter six, gene expression in SDHD, SDHAF2 (formerly known as the PGL2 locus) and sporadic head and neck paragangliomas is investigated. An attempt is made to distinguish these genetic subgroups on the basis of their gene expression profile and to link mutations in SDHD and SDHAF2 to specific tumor behaviour and tumorigenic pathways.

Chapter seven consist of a general discussion on hereditary paragangliomas in the Netherlands