A number of inflammatory, infectious, neoplastic and idiopathic disorders affect the eye and the central nervous system (CNS) concurrently or at different time frames. These conditions pose a diagnostic challenge to the clinician since they may present with similar ocular and neurological manifestations. The purpose of this review is to describe major neurological syndromes including multiple sclerosis, Vogt-Koyanagi-Harada disease, other autoimmune syndromes, and several infectious diseases which may affect the eye. This article may serve as a guide for the diagnosis and treatment of such disorders. It should be noted that these conditions have been viewed from a neurologist’s perspective thereby neurologic involvement is stressed. The frequent concurrence of uveitis and central nervous system (CNS) diseases may be derived from the common embryogenic developmental pathway of the posterior segment and the CNS. Analogies between the blood brain barrier and the blood retinal barrier, and “immune privilege” involved in the prevention and blockage of noxious immune system reactions both in the eye and CNS also reflect the common origin of these organs. A North-American reviewMany CNS disorders affect the posterior segment of the eye. Despite their heterogeneous origin , upon which the most recent classification is based, these diseases have a single functional unit as their target, represented by the CNS and the eye.Such pathogenetic associations may underlie variability in prevalence related to ethnicity. For instance Behçet disease, which is rare in the USA (0.3% of uveitic entities)Apart from these considerations, we also have to point out that, the most represented patients in Smith’s work was a group of people suffering from CNS disease of unknown origin (meningitis and myelitis) or peripheral nervous system (PNS) disease (angiitis).1From an etiological point of view, the two broadest and most recent surveys from the USA This review will discuss major entities which present with neurological and ocular inflammatory diseases.The following classification is a modification based on the work by Brazis et al.5Demyelinating diseases- Multiple sclerosis- Monosymptomatic optic neuritisInfectious diseases- Viral , acquired immunodeficiency syndrome (AIDS), subacute sclerosing panencephalitis (SSPE)- Bacterial - Mycobacterial - Spirochetal - Protozoal - Nematodal or parasitic (gnathostomiasis)Inflammatory or Autoimmune- Polyarteritis nodosa- Wegener granulomatosis- Rheumatoid arthritis and Sjögren’s syndrome- Systemic lupus erythematosus- Sarcoidosis- Behçet’s disease- VKH disease- APMPPE Neoplastic- Lymphoma- Paraneoplastic syndromesMultiple sclerosis (MS) may affect the entire CNS and must be distinguished from monosymptomatic optic neuritis (MSON) which presents with ocular symptoms but no systemic involvement. MSON can cause a single episode, or lead to multiple recurrences of ocular involvement. A number of patients, after some years of quiescence, evolve to MS such that 40% of MSON cases develop MS in 8 to 10 years .Isolated or combined with other neurological symptoms, optic neuritis mainly affects young patients (less than 30 years of age) and is rarely seen after the age of 45. In the latter case, the disease presents with medullary deficits and a rapidly disabling course. Based on such multiform clinical findings, one may assume that MS is not a single clinical entity, but a group of demyelinating diseases which can be differentiated from each other not only on the basis of symptoms and course, but also according to etiology, genetics, immunopathogensis and response to therapy.Optic neuritis usually presents with acute unilateral blurring of vision, transient photopsia, ocular or orbital pain (due to optic nerve edema) and other neurological complaints including sensory, motor, cerebellar and urinary bladder symptoms. Other symptoms such as diplopia, vertigo (due to proximity of the oculomotor area to the equilibrium area), trigeminal neuralgia, neuro-vegetative deficits (mainly orthostatic hypotension) and cognitive deficits are infrequent.The course of MS is characterized by relapses and remissions (more commonly during the early stages of the disease) but may occur as a primary or secondary chronic-progressive disease. In later stages, a number of variations and mixed forms such as chronic-progressive disease with overlapping relapses are possible.Magnetic resonance imaging (MRI) shows hyperintense round/oval demyelinating lesions mainly in periventricular areas on T2-weighted sequences . If the A classic acute attack of MS or MSON usually lasts more than 24 hours; if visual or non-visual symptoms are transient, they represent only brief deterioration due to CNS causes leading to nerve conduction delay. In these cases symptoms rapidly disappear when the exciting cause subsides.Intermediate uveitis is associated with MS; 10–20% of uveitic subjects show MS symptoms after some years, and a similar number of MS cases present with major anatomo-pathological signs of intermediate uveitis. These may include retinal peri-vasculitis mainly affecting veins in the active phase, vascular sheathing in the chronic phase, and perivenous sclerosis following retinal vasculitis in over 30% of chronic-progressive MS patients  types 1 and 2, varicella zoster virus (VZV), cytomegalovirus (CMV), and Ebstein-Barr virus (EBV), are diseases that typically affect immunocompromised, HIV-positive, atopic, or transplanted individuals or patients receiving cytostatic, steroid or immune-suppressive therapy. However, an increasing incidence is noted nowadays in healthy people. The status of the host’s immune system defines the outcome of these infections and complications related to these viral entities.Ocular involvement presents in two major forms: 1) anterior uveitis and 2) viral retinopathies.ARN is usually caused by VZV and HSV, but more rarely by CMV12All herpetic infections show tropism for sensory ganglia inside which the virus remains dormant during periods of latency. This mechanism explains how herpes viruses spread to ganglia-derived areas (dermatomes).Neurological manifestations are usually not related to immune system dysregulation or suppression. HSV neurological infections cause: 1) encephalitis and 2) recurrent aseptic meningitis., frontal and temporal lobe lesions respectively . Necrotizing acute encephalitis is frequently localized to the temporal lobes, sometimes with meningeal involvement, and presents with typical symptoms of meningoencephalitis , associated with memory, personality and speech disturbances because of hippocampalectively .13,14VZV infection typically presents with sensory ganglia involvement including thoracic or cranial  ganglia with dermatomal cutaneous manifestations. In the central nervous system, VZV produces herpes zoster ophthalmicus which can involve the eye where it produces anterior granulomatous uveitis often resulting in sectorial iris atrophy. Although herpes zoster is due to reactivation of dormant varicella virus, it is an acute viral infection that requires prompt treatment with systemic antivirals to avoid complications which occurred in the era before antivirals were available.Apart from these classical forms, a new neurological entity called “Zoster sine herpete” has been described, thanks to the recent introduction of polymerase chain reaction (PCR) tests. It presents as aseptic meningitis or encephalitis (with cerebellar tropism), sometimes as transverse myelitis, or cerebral vasculopathy derived from granulomatous angiitis (late CNS vasculitis) with deep ischemia of the basal ganglia, temporal lobes, and the internal capsule.9Primary herpetic infection results in axoplasmic transport of the virus to sensory ganglia where it establishes latency. Reactivation leads to retrograde virus transport. The true path by which the virus reaches the CNS has not been established, one possibility is that the reactivated virus located in ganglia may spread to the anterior and middle cranial fossa.The mechanism by which herpes viruses affect the eye is the double occurrence of a lytic reaction followed by intraocular inflammation.12History and clinical examination of herpetic infections of the CNS and eye are usually diagnostic because they are well-defined entities. In controversial or atypical presentations, molecular diagnosis of cerebrospinal or ocular fluids by PCR provides high sensitivity (96%) and specificity (99%)For diagnostic purposes in equivocal cases of meningoencephalitis, the following techniques may be useful: 1) cerebrospinal fluid (CSF) analysis revealing typical features of monocytic pleocytosis, high protein content and normal glucose content; 2) neuro-imaging studies including computed tomography (CT) or ideally MRI with characteristic findings of symmetric necrosis of the temporal lobes; 3) electroencephalography typically showing periodic spike waves on temporal derivations. These investigations may facilitate diagnosis leading to prompt and specific therapy with systemic antiviral agents, preventing severe ocular and neurological sequelae and sometimes death.13Recurrent aseptic meningitis is mainly due to HSV 2; history is positive for herpes genitalis and this infection seems to spread via the oculogenital route to the eye and via sacral sensory ganglia to the CNS. It presents with symptomatic meningitis and fever, meningism, an intense headache lasting for some days followed by alternating recovery with symptomatic periods. Typical CSF findings are lymphocytic pleocytosis and high protein levels. PCR investigation is almost always positive; viral culture is positive only in the early stages of the disease.CNS and ocular infections due to CMV have been reported following congenital infections and in patients with AIDS in the pre-HAART  era due to severe immunodeficiency, sometimes causing death. Currently, the course of CMV retinitis or meningoencephalitis in immunocompromised hosts has dramatically changed and CMV infection may present as the first and only clinical manifestation of AIDS.CMV infection in the eye presents as active retinitis with a typical “pizza-like” appearance consisting of white-yellowish cotton wool areas of necrotizing retinitis associated with hemorrhage and vascular sheathing with mild vitritis evolving towards progression of retinitis and retinal scarring. Disk atrophy follows papillitis or acute anterior optic neuritis in the acute phase CMV encephalic infection is mainly localized to the brainstem (similar to Behçet’s disease). Typical symptoms are lid ptosis, oculomotor disturbances and nystagmus. Ventriculitis and inflammatory poly-neuropathies are infrequent.Bell’s palsy may be caused by any of the viral diseases under this category; however, to date, there has been no association with uveitic entities.HTLV-1 is the cause of T-cell leukemia and of a peculiar myelopathy  which may be associated with ocular manifestations including keratoconjunctivitis sicca, corneal lesions, retinal vasculitis and uveitis, otherwise known as HTLV-1 associated uveitis or HAU. It is more frequent in Japanese, African, Caribbean and South-American patients.2 hyperintense areas related to demyelinization.Indirect AIDS consequences are opportunistic infections and tumors. HIV-1 viral infection can directly induce a number of neurological syndromesOcular manifestations of AIDS can be categorized into four main groupsWest Nile virus (WNV) infection is a mosquito-borne disease widely distributed in all continents. Since 1999, when the first outbreak of WNV encephalitis occurred in New York city, it has had an impressive spread, representing a recent example of geographical evolution of a disease formerly restricted to specific areas.20WNV, similar to other arbovirus (flavivirus family) transmitted infections such as dengue, yellow fever and Japanese encephalitis, shares the common feature of neuro-invasiveness with grey matter affinity, with loci of involvement in the cortex, basal ganglia, brainstem, cerebellum and spinal cord. Less than 1% of infected individuals develop severe neuro-invasive diseases which can be classified into three major but sometimes overlapping, clinical syndromes: meningitis, encephalitis and acute flaccid paralysis.21Meningitis has the usual manifestations: fever, headache and neck stiffness. Encephalitis shows focal symptoms related to the site of involvement such as epilepsy (cortex), tremor and involuntary movements , parkinsonism (substantia nigra) and ataxia (cerebellum). Encephalitis incurs severe sequelae and a high incidence of death. Acute asymmetric flaccid paralysis, similar to poliomyelitis , has an unfavourable outcome with variable recovery.Before the onset of ocular or CNS symptoms, many patients are healthy without any signs of immunosuppression, while some (approximately 20%) complain of mild flu-like symptoms including headache, fever, malaise, gastrointestinal upset, skin rash, neck rigidity and changes in mental status.Ocular manifestations are uncommon although chorioretinitis may be more frequent than previously reported.Detection of WNV–specific IgM in the serum or CSF provides strong evidence of recent infection. IgM usually becomes detectable 8 days after the onset but may become elevated after some monthsSubacute sclerosing panencephalitis (SSPE) is an encephalopathy of subacute course affecting children and adolescents caused by an aberrant measles virus called the SSPE virus. It is characterized by progressive encephalopathy involving both grey and white matters with a fatal outcome in most cases.23Ocular involvement is seen in more than half of the cases and is usually bilateral. The macular area is usually involved by retinitis or pigmentary changes. Optic neuritis is present in two-thirds of cases with ocular involvement. Ocular findings may sometimes precede CNS signs and symptoms. Macular involvement may resemble acute posterior multifocal placoid pigment epitheliopathy (APMPPE) which is one of the differential diagnoses.24Ocular involvement in subjects affected with syphilis is strongly suggestive of CNS involvement and should be considered synonymous with neuro-syphilis.From an ophthalmological point of view,Acute meningitis can occur in 1 to 2% of patients with secondary syphilis, and can cause papilledema due to a rise in intracranial pressure. With papillitis and vitritis, patients complain of acute visual loss; neuroretinitis manifests with hard exudates in the peripapillary area. Another manifestation is optic “perineuritis” which may be suspected in a subject with optic nerve head swelling without ocular symptoms which is due to mild inflammation of meningeal sheaths of the optic nerve. Many patients with ocular signs do not show systemic evidence of the syphilitic disease.Ocular manifestations also include anterior (granulomatous and non-granulomatous) uveitis, intermediate uveitis, panuveitis, chorioretinitis, retinal vasculitis, and necrotizing retinitis; the latter may be the first presentation in patients with HIV infection. Ocular syphilis has to be treated exactly in the same way as neuro-syphilis.Tertiary syphilis may present with meningovascular and late parenchymal consequences and from an ophthalmological point of view, with interstitial keratitis or different forms of uveitis. The slower and more chronic course of tertiary syphilis is the only difference between secondary and tertiary stages of the disease. A small number (5%) of tertiary neuro-syphilis patients suffer from optic atrophy.The protean expressions of syphilis, particularly ocular involvement, have caused the disease to be designated as “the great imitator”.Lyme disease is a tick-borne infection due to different types of Borreliae, called “the new great imitator” since it displays a broad variety of systemic symptoms ranging from skin erythema migrans to neurologic or cardiac lesions. Ocular manifestationsNeuro-ophthalmic manifestationsCranial neuropathies may result from direct infection, autoimmune inflammatory response (i.e. vasculitis of vasa nervorum), or raised intracranial pressure. Trigeminal nerve involvement may present as bilateral decrease in corneal sensation or as ocular and periocular pain. Papilledema may occur with meningitis and increased intracranial pressure. Horner’s syndrome, tonic or dilated pupils, and photophobia have also been described as signs of encephalopathy or meningitis.2 weighted images). In demyelinating disease, involvement of basal ganglia grey matter is absent.30In the third stage, circumscribed vasculitic brain lesions appear and result in different symptoms according to the site of involvement  and an MRI pattern similar to that of MS  is a multi-systemic infection with predominantly gastrointestinal manifestations caused by an actinomyces (Tropheryma Whippelii). Major presenting symptoms include diarrhea, impaired intestinal absorption and progressive weight loss. Once the organism enters the bloodstream via the intestinal lamina propria, ‘colonization’ of the CNS occurs in a variable number of cases (43–100%) according to differing studies. Asymptomatic phases alternate with sudden relapses, suggesting direct action of the microorganismOcular manifestations are secondary to CNS involvement. According to a large reviewCat scratch disease is a systemic zoonosis caused by a gram-negative bacillus (Bartonella henselae). Systemic infection after primary inoculation is associated with localized erythema and regional lymphadenitis with flu-like symptoms.The major neurological manifestation (in 1–2% of cases) is encephalopathy from infective micro-localizations, involving both hemispheres and underlying cerebral structures. Characteristic symptoms are epileptic seizuresThe largest reported case seriesWhen the eye is involved, the disease is characterized by Parinaud’s oculoglandular syndrome including red eye, mucoid conjunctival exudates and unilateral lymphadenopathy. After hematogenous spread neuroretinitis, chorioretinal lesions , and sometimes capillary angiomatous proliferation may be noted.37Gnathostoma , which is endemic in subtropical Asia (from Thailand to Japan) and in some parts of the American continent . In the “visceral” stage of its complicated life cycle which follows the “cutaneous” stage, this parasite can involve all internal organs, including the eye and CNS. Neurological manifestations are associated with a high percentage of mortality and include (in order of appearance): radiculo-myelitis, radiculo-myelo-encephalitis, eosinophilic meningitis and subarachnoidal hemorrhage (SAH). Parasites enter the CNS through nerve roots  and give rise to symptoms (e.g. radicular pain), which persist for some days. Afterwards, limb paralysis  can occur when parasites ascend to the brain. Finally, eosinophilic meningitis and SAH, the most frequent causes of death, ensue. In Thailand, 6% of all SAH in adults and 18% of SAH in children appear to be due to gnathostomiasis.Gnathostomiasis is a parasitosis due to nematodes of the genus The eye is the only organ where gnathostoma can be visualized, usually in the anterior chamber, but also rarely in the vitreous. Ocular symptoms include uveitis (mostly anterior), iritis, intraocular hemorrhage, glaucoma, retinal scarring and retinal detachment.Because of the “immune privilege” status of the eye, eosinophilia is less marked with ocular disease than when other organs are involved.38Anterior uveitis can occur in meningococcal sepsis, a potentially fatal illness and less frequently with meningococcal meningitis.The most frequent neurological manifestation of tuberculosis is meningitis, which unlike other causes of bacterial meningitis is characterized by clear CSF with monocytic pleocytosis due to rupture of sub-ependymal tubercles. Intra-parenchymal granulomas can induce epilepsy. Other symptoms of neuro-tuberculosis are papilledema, retrobulbar optic neuritis and oculomotor nerve palsy. Ocular involvement is polymorphic and any form of uveitis is possible, the most common being posterior uveitis, serpiginous-like choroiditis and subretinal abscesses due to foci of caseous necrosis.Mycoplasma pneumoniae infection can cause aseptic meningitis. Symptoms related to involvement of the left hemisphere, i.e. right hemiparesis and motor aphasia, have been described by Perez and FosterNeurological involvement in cryptococcosis includes meningitis , peri-optic meningeal infiltration leading to retrobulbar optic neuritis and cranial nerve (mostly sixth nerve) palsy, and parenchymal localization causing dementia. In 40% of cases of neuro-cryptococcosis, ocular involvement is present, mostly as multifocal chorioretinitis.Leprosy affects the peripheral nervous system causing mainly sensory poly-neuropathy with neuropathic pain and paresthesia in the first stage, and sensory loss with susceptibility to traumatic limb injuries in more advanced stages. Acute or chronic anterior uveitis may be present in 10–50% of cases.19Cerebral vasculitides (CVs) are associated with retinal vasculitis and uveitis in different percentages. CVs are present in a number of systemic autoimmune disorders and show common clinical aspects: 1) a symptomatic pattern including headaches , multifocal deficits (secondary to ischemic lesions) and neuropsychiatric symptoms ; 2) blood signs of acute inflammation; 3) coexistence of central and peripheral neurological deficits ; 4) frequent extra-neurological involvement, because vasculitides are systemic diseases.On MRI vasculitic lesions are usually hyperintense on T2- but sometimes also on T1-weighted sequences . If an iCVs which commonly affect the eye include polyarteritis nodosa, Wegener’s granulomatosis, rheumatoid arthritis  and systemic lupus erythematosus.Polyarteritis nodosa (PAN) is a necrotizing segmental inflammation of small and middle sized arteries; it is frequently (70%) associated with hepatitis B virus (HBV) seropositivity and clinically presents as peripheral involvement (mononeuritis multiplex in 70% of cases) as compared to CNS lesions (ischemic or hemorrhagic stroke in 10%). Vasculitic involvement can affect all ocular structures leading to episcleritis, diffuse/nodular scleritis, vasculitic or occlusive retinopathy, choroidal vasculitis (the most frequent manifestation), ischemic optic neuropathy and orbital vasculitis (with extraocular muscle ischemia and amaurosis fugax).39Wegener’s granulomatosis (WG), together with Churg-Strauss arteritis and microscopic polyangiitis, is one of the ANCA (anti-neutrophil cytoplasmic antibody)-positive vasculitides. Neurological manifestations are caused by invasion from paranasal sinuses to the cranial nerves causing oculomotor nerve deficitsRheumatoid arthritis (RA) can be the cause of central or, more frequently peripheral (vasa nervorum) vasculitic involvement; however, retinal vasculitis is rare. Sjögren’s syndrome, an independent or RA-linked disease which was once thought to be a variant of RA, presents with frequent, both central and peripheral neurological symptoms which precede the diagnosis of the condition in most cases.Systemic lupus erythematosus (SLE), one of the most polymorphous diseases, can cause neurological manifestations with different mechanisms. Focal symptoms  are due to circumscribed lesions of vasculitic nature . At onseNeuropsychiatric disorders  are caused by direct action of anti-neuron antibodies including anti-P  and anti-N  antibodies.43As in PAN and WG, any ocular structure can be affected, from the lacrimal system to the retina. Inflammatory arteriolar vasculopathy is more severe in the presence of anti-cardiolipin antibodies and is often complicated by proliferative retinopathy in the case of major vascular occlusion, by pigmentary alterations (“pseudoretinitis pigmentosa”), or by exudative retinal detachment (secondary to choroiditis) although less common than retinal involvement.44Sarcoidosis is a protean multisystemic disease of unknown origin caused by dysfunctional T-lymphocytes, phagocytes, and epithelioid cells generating non-caseating granulomatous multi-organ inflammation. Neurological and ocular manifestations of sarcoidosis are uncommon and present in 5 to 10% of cases according to different reports. No predisposing factors are evident. Neuro-sarcoidosis (NS) begins in the leptomeninges and spreads through the subarachnoid space to other meningeal structures, the hypothalamus, hypophysis, chiasma and cranial nerves, and through perivascular areas towards encephalic parenchyma. Infrequently, encephalic vasculitis may be present.Meningeal localization is evident in more than 90% of autopsies but clinical manifestations of such involvement are less frequent (8–64%).According to different studies,Optic neuritis (present in 8–13% of symptomatic patients) is caused by granulomatous optic nerve infiltration, intracranial hypertension following meningitis, extrinsic compression, and vasa nervorum vasculitis. Related symptoms are pupillary abnormalities (tonic pupils) and hemianopsia .Single or multiple cranial nerve involvement caused by nerve infiltration, vasa nervorum vasculitis, intracranial hypertension or compression. The seventh cranial nerve is mainly involved (20–50%) because of parotid compression, followed by oculomotor manifestations (diplopia), the so-called Parinaud’s syndrome.Peripheral nerve sensory-motor neuropathy.Cerebral localization .Granulomatous infiltration myopathy.Neuroendocrine symptoms (15–23%) including hypogonadism and diabetes insipidus because of granulomatous involvement of the hypothalamus and hypophysis.Polymorphic ocular manifestations of sarcoidosis may coexist; these include adnexal involvement , conjunctivitis, episcleritis, scleritis, keratitis, granulomatous or non-granulomatous anterior uveitis, posterior uveitis , optic nerve involvement . Uveitis is the most frequent sign in NSOcular manifestations in subjects suffering from NS are more common than previously thought. Patients with neurological symptoms and associated intraocular inflammation should undergo work-up for sarcoidosis including brain and orbital MRI  and 10 aOral steroids are the mainstay of treatment for NS and associated intraocular involvement; when they fail, systemic immunosuppressive or biological agents can also be used to avoid chronic sequelae of the disease.Behçet’s Disease (BD) can be defined as a systemic vasculitis that usually affects venules, sometimes veins, rarely arterioles and exceptionally arteries. These different frequencies remain unexplained as yet. The eye is the most commonly involved organ, with typical manifestations of relapsing-remitting panuveitis and retinal vasculitis. As a systemic disease, BD may also affect the CNS.Neuro-Behçet (N-BD) shows ethnic variations and also disparity within the same ethnic group and from one study to another; this depends on inclusion criteria and characteristics employed in each study. The following illustrates this variability. In a group of 558 Turkish BD subjects Akman-DemirThe frequency of BD neurological manifestations varies from 3% to 30% and seems to be high among Arabs, intermediate in Turkish subjects and low in Japanese individuals.64If small veins are involved, the condition corresponds to the meningo-parenchymatous form (MP).If veins are affected, the term angio- BD is employed.If small arteries (vasa vasorum) are involved, aneurysms may rarely develop.MP is the prevalent form and the MP/ angio-BD ratio varies from 3/1 to 11/1 according to different statistics. Male subjects are more frequently affected by BD and N-BD. The pathogenesis of MP and angio-BD is an unsolved problem; we only know that the two forms have different pathogenic mechanisms, in fact they rarely appear in the same patient.MP shows tropism for the brainstem in which the venous circle appears to have slower blood flow than other districts; however, only the mesencephalon and pons are involved whereas the bulbus remains undamaged. Other preferred CNS structures are the cerebellum, basal ganglia, internal capsule and meninges.The course of MP is very similar to MS, with relapsing-remitting primary and secondary chronic-progressive forms; the only difference is in the sudden onset of symptoms and the lower frequency of MP relapses. As a relapse symptom, headaches are very frequent, are mostly hemicranial (with or without aura) rather than meningeal, and are related to lesions in the periaqueductal grey matter (a pontine structure involved in the pathogenesis of migraine).Bilateral pyramidal signs are the most common symptoms of MP; other symptoms include behavioral disorders  and less frequently brainstem signs, e.g. several forms of ophthalmoplegia . Meningeal symptoms are rare despite the specific “meningo-parenchymatous” involvement, optic neuritis is also rare.Intracranial hypertension is the main presentation of angio-BD and is due to thrombosis of venous sinuses  where flow is low. Intracranial hypertension gives rise to papilledema and often abducens palsy from extrinsic compression. Headaches are very frequent in the acute phase and are sometimes associated with fever.When compared to MP, angio-BD shows an earlier outbreak, an invariably acute course (it is never chronic-progressive), a better response to therapy and a more favorable prognosis.Rare forms of N-BD are: aneurysms, indirectly caused by vasa vasorum vasculitis and by sectorial arterial wall ischemia ; pseudotumor , characterised by bulky localisation of the MP form; different types of neuropathies  disease is one of the most common uveitic-related neurological diseases. This condition affects other apparently heterogeneous tissues including the leptomeninges, cochlea and skin which are similar in the presence of melanocytes. In fact, during embryogenesis, most pigment cells arise from cranial or truncal portion of the neural crest and migrate to the skin, hair bulbs, choroid, inner ear, leptomeninges and other tissues.4 ad HLA-DR53 .which ;re predominant in some ethnic groups  and to the HLA-DRB 1*0405 allele, which is predominant in Brazilians VKH is related to HLA-DRThe incidence of neuro-otologic symptoms is related to ethnicity. Meningism and the CSF pattern of monocytic meningitis are prevalent in the Japanese. Otological symptoms and perceptive deafness are more frequent in the Japanese, whereas tinnitus shows a similar distribution in all studied populations apart from Hispanic populations who are less affected by otological symptoms. Hearing involvement can persist over many years.The meninges are affected in the prodromal stage which is characterized by fever, headache, neck stiffness and psychomotor retardation together with cochlear involvement which causes dysacusis and tinnitus. Neuro-otologic symptoms are more prominent in the second stage, when uveitis also occurs (i.e. the acute uveitic or uveitic-acoustic stage). Vestibular symptoms are rare, probably because different types of melanocytes are present in the inner ear. The skin is involved in the third stage, i.e. the convalescent stage, when “sunset glow fundus” and retinal pigmentary abnormalities can occur. In the fourth stage, if the disease is untreated, ocular complications can appear.72In the acute stage, fundus findings consist of bilateral multifocal serous retinal detachments in the posterior pole together with optic nerve inflammatory edema. FA reveals retinal pigment epithelium (RPE) hyperfluorescent dots, multiple round areas of dye pooling in the subretinal space, and optic disc leakage. Indocyanine green angiography (ICGA) demonstrates patchy filling delay of choroidal vessels and the choriocapillaris, diffuse dye leakage, and multiple hypofluorescent spots. Optical coherence tomography (OCT) shows large serous retinal detachments in the central area.In the convalescent stage, fundus examination reveals the typical sunset-glow appearance with irregular linear pigmentation in the posterior pole. FA shows granular hyperfluorescence reflecting RPE damage and ICGA demonstrates significant improvement in filling delay, but hypofluorescent dark spots  persist for a long period.80In early stages, neurological manifestations appear before ocular symptoms. Main symptoms are fever, headache, nausea, photophobia, meningism and neck stiffness, orbital pain (related to optic nerve edema or hyperemia) and galea capitis hyperesthesia. A recent study identified anterior ischemic optic neuropathy (AION) with irreversible visual field defects (6 cases in 52 consecutive patients).753) causing confusion with an infectious process. A recent review by Andreoli and FosterCSF examination allows early diagnosis by showing a not very high monocytic pleocytosis  is a primary inflammatory choriocapillaropathy often provoked or preceded by a viral infection. The condition may present with neurological symptoms which are variable or can follow ocular manifestations. Typical ocular symptoms are photopsia, scotoma and visual loss. Ocular findings include deep bilateral areas of yellowish discoloration in the posterior pole with multiple serous exudative retinal detachments, together with mild vitritis and anterior uveitis. ICGA shows round geographic hypofluorescent areas which represent choriocapillaris non-perfusion .The most frequent neurological symptoms is meningism with lymphocytic pleocytosis and headaches.APMPPE must be suspected in a young (20 to 30 year old) patient with a viral flu-like syndrome, stroke of unclear etiology, meningism with CSF pleocytosis, and ocular symptoms such as photopsia, scotoma and visual blurring.Kawasaki’s disease or the “mucocutaneous lymph node syndrome” can present as a bilateral anterior uveitis with or without aseptic meningitis and monocytic pleocytosis in children and young adults.Crohn’s disease and ulcerative colitis can cause ocular signs including iritis, conjunctivitis, episcleritis, scleritis and uveitis; and inflammatory/ischemic neurological symptoms i.e. optic neuritis and other cranial nerve neuropathies, especially oculomotor nerves.Cogan’s syndrome consists of acute interstitial keratitis and vestibulo-auditory dysfunction. The latter deficit may also occur in Eale’s disease, which is very frequent in India; the main sign is retinal periphlebitis without true uveitis.Relapsing polychondritis is an autoimmune disease targeting type II collagen characterized by destructive and relapsing inflammation of the cartilage in organs such as the nose, earlobe and trachea. Ocular manifestations include episcleritis, scleritis and iridocyclitis; neurological symptoms are encephalic . Cranial nerves, chiefly the optic nerve and sensorineural deafness, are involved in necrotizing vasculitis Susac’s syndrome (retino-cochleo-cerebro vasculopathy) is a very rare immune-mediated microangiopathy characterized by encephalopathy, branch retinal arterial occlusion (BRAO) and hearing loss of unknown etiology. The main presenting symptoms are CNS features.82Presenting symptoms of encephalitis are personality and behavioral changes, associated with intense headaches and migraine. MRI shows small multifocal supra-tentorial lesions  mainly involving the white matter, however, the grey matter is sometimes affected as well.Visual symptoms are related to the site of occlusion: if in the retinal periphery  there are no visual complaints; if it involves the central area, the affected subject may complain of visual impairment which is usually bilateral and may sometimes be the presenting symptom. Branch retinal artery occusion may be extensive or limited and FA is the main imaging technique to show it; after resolution silver streaks are noted which replace the occluded arterioles. Cerebral and visual symptoms are joined to some hearing loss forms.83Differential diagnoses include MS and ADEM . The differentiating feature on MRI is that the lesions related to the two latter conditions are located inside the undersurface of the corpus callosum and not in the central area as in Susac’s syndrome.Susac’s syndrome has a self-limiting but long and fluctuating course, ranging from one to five years with tendency towards spontaneous improvement.Although the etiology is unknown, findings from brain biopsies suggest small vessel occlusive vasculitis caused by an autoimmune, viral-mediated or hypercoagulability disease. In this way, ocular, brain and cochlear arteriolar micro-vascular occlusion are explained.85No treatment is evidence-based because the disease is very rare. However, high doses of intravenous steroids and immuno-suppressive agents are considered to be the mainstay of therapy.The eye can be involved in both systemic and diffused hemoblastomas, particularly in B small-cell non-Hodgkin lymphoma (NHL), and in a specific form of lymphoma, primary CNS lymphoma (PCNSL) which is more appropriately defined as CNS and ocular lymphoma i.e. a single neoplastic manifestation that can be localized to the CNS, the eye, or both.PCNSL represents 4–6% of all cases of primary CNS lymphomas and 1–2% of extranodal lymphomas.According to a an older report, ophthalmic localizations precede neurological onesWhen ocular symptoms  occur prior to neuro-radiologic manifestations, the disease is designated primary intraocular lymphoma (PIOL). Infrequently, lymphoma is localized to the eye or the brain, probably due to diagnostic inaccuracy.Typically, in a female patient older than fifty, chronic uveitis without a well-defined etiology which lacks classic signs of anterior segment  or retinal inflammation (vasculitis) and is unresponsive to steroids, should raise the suspicion of lymphoma. Certain infectious agents including HSV-8, EBV and toxoplasma gondii have been suggested in the pathogenesis of this condition by stimulating B lymphocytes to evolve into lymphomatous cells.93The two main pathogenic hypothesesFrom an immunogenetic point of view, the majority (60–75%) of PCNSL cases demonstrate a deletionIn cerebral PCNSL, the lesions are usually supratentorial, multicentric and localized in the basal ganglia, corpus callosum and periventricular areas with a predilection for the frontal lobe.96The prevalence of frontal lobe involvement corresponds to personality and sleep disordersOcular localization is rare but is the most common malignancy causing a “masquerade” syndrome. PIOL is a diffuse large B-cell lymphoma2 cytokine group and acts as a “growth factor” for B-cells and therefore contributes to the development and proliferation of PCNSL by inhibiting Th1 cytokines and consequently, T cell cytotoxic effects against malignant B-cells. Furthermore, it increases BCL-2 gene expression on B-cells and prevents their apoptosis. IL-10 is increased in PCNSL serum (similar to AIDS-associated lymphomas and EBV-related Hodgkin’s disease) and CSF. In the aqueous humor, an increased value (more than 1.0) of IL-10 and IL-10/IL-6 (a cytokine mainly produced by inflammatory cells) ratio is diagnostic for the presence of PIOL/PSNCL. In the serum, this ratio is related to both the clinical expression and the prognosis of the disease.Another very useful diagnostic tool is the detection of interleukin-10 (IL-10) in the serum and aqueous humor produced by malignant B-cellsEye and brain CT are usually sufficient to make a diagnosis of PCNSL but a specific CT or MRI pattern does not exist. Some features are frequently presentTypically, lesions involve the corpus callosum, basal nuclei, or periventricular white matter1 hypointense and T2 hyperintense patterns, similar to MS.MRI without contrast shows PCNSL localizations as TParaneoplastic optic neuropathy (PON), has been described in cases of microcytic lung carcinoma and thymomaAs highlighted in this review, a number of disorders with heterogeneous origins present with oculocerebral symptoms and signs, with onset sometimes in one or both organs. These syndromes may be difficult to manage because they are uncommon and difficult to diagnose. Treatment requires qualified interdisciplinary knowledge and management at specialized referral centers.


Radiation therapy is an established modality in the treatment of head and neck cancer patients. Compromised wound healing in irradiated tissues is a common and challenging clinical problem. The pathophysiology and underlying cellular mechanisms including the complex interaction of cytokines and growth factors are still not understood completely. In this review, the current state of research regarding the pathomechanisms of compromised wound healing in irradiated tissues is presented. Current and possible future treatment strategies are critically reviewed. Radiation is employed as neoadjuvant, primary and adjuvant therapy for head and neck cancer. Complications after radiation therapy occur in up to 60 percent of surgical patients. Clinical sequelae include skin atrophy, soft tissue fibrosis, desquamation, epithelial ulceration, fistula formation and major vessel rupture,2. ImpaiAdequate wound healing involves interactions of cells. Cell biologic mechanisms relevant to the process include interaction of keratinocytes, fibroblasts and endothelial cells. EpithelHemostasis and inflammation , are followed by proliferation  and maturation ,11. ThesWound healing occurs in an ordered sequence of cellular interactions. Repetitive radiation injury disrupts this highly organized sequence of events, resulting in repetitive inflammatory responses and ongoing cellular regeneration.There is an important distinction to be made between the early and the late side-effects of radiation therapy: Early side effects include erythema, dry desquamation, hyperpigmentation and hair loss. Late efThe inflammatory and proliferative phases may be disrupted by the early effects of radiation. Affected factors during the inflammatory phase include transforming growth factor beta (TGFβ), vascular endothelial growth factor (VEGF), tumor necrosis factor-α (TNF-α), interferon-γ (IFN- γ) and proinflammatory cytokines such as interleukin-1 and interleukin-8. These cThe proliferative phase is characterized by granulation tissue formation, re-epitheliaziation and neovascularization. This phase is mainly regulated by TGFβ, VEGF, epidermal growth factor (EGF), fibroblast growth factor (FGF) and platelet-derived growth factor (PDGF). Nitric During the remodeling phase, matrix metalloproteinases (MMP) and their tissue inhibitors are central to the process,19. MMP-Keratinocytes represent a crucial cell type in the repair of late epithelial wounds and ulcers. Multiple molecular biological changes are observed in this cell after radiation when compared to radiation-naïve skin. In human radiogenic wounds, these cells show a shift in the expression from high molecular keratins 1 and 10 to the low molecular keratins 5 and 14. In non-healing ulcers, keratinocytes display a decreased expression of transforming growth factor-alpha and –beta(1), fibroblast growth factor 1 and 2, keratinocyte growth factor, vascular endothelial growth factor, and hepatocyte growth factor. Expression of the matrix metalloproteinases 2, 12 and 13 has been shown to be elevated in irradiated human keratinocytes and fibroblasts.Fibroblasts play the central role in wound healing through deposition and remodeling of collagen fibers. In irradiated tissue, fibroblasts have been shown to generate a disorganized deposition of collagen bundles. One likely mechanism resulting in disorganized collagen deposition is dysregulation of MMP and TIMP. These enzymes regulate extracellular matrix synthesis,22. As TEstablished strategies in treating radiogenic ulcers with delayed and inadequate healing include standard wound care, vacuum-assisted devices, substitution of nutritional deficiencies, and measures to optimize blood and oxygen supply,23.Hyperbaric oxygen seems to optimize the partial pressure of tissue oxygen. It is frequently used for the treatment of osteoradionecrosis-27. The Currently, hyperbaric oxygen therapy is clinically used in chronic diabetic ulcers and wound healing complications after radiotherapy. Its efficacy has been proven by several randomized trials, but it is it is important to emphasize that for both diabetic wounds and radiation injuries, this therapy is used in conjunction with standard wound care management techniques. A possiResearch into new therapeutic approaches to treat radiogenic ulcers includes: Special dressings, injection of (multipotent) cells, topical administration of active substances and the use of growth factors Table.Different types of membranes and other covers of radiogenic skin injuries have been developed in the past.Hydrogel membranes have a stable, flexible and transparent structure. Data of Lu et al. indicate that the hydrogel membranes act as a semiocclusive dressing and maintain a moist environment over the wound bed. Through this mechanism, re-epithelialization is thought to be enhanced because of the accumulation of cytokines and growth factors that support wound healing. The useBacterial cellulose impregnated membranes and poviargol, an antiseptic substance containing silver, was shown to accelerate the healing of irradiated wounds in an animal model. A recenA human deceased-donor skin allograft provides dermoprotection and promotion of re-epithelialization. It is utilized until autografting is possible or re-harvesting of donor sites becomes available. This trWound healing is a complex mechanism that requires active cellular interactions. These interactions are only possible if a sufficient number of intact and healthy cells are present. Radiation leads to impaired cellular activity and cell death. Injection of cells has the potential of enhancing wound healing in irradiated tissues. Nevertheless, there is a significant risk of infection and possible tumor induction by injecting viable cells in vivo. All of the studies discussed below have to be seen in this context. All interventions are experimental and presently not applied in routine clinical practice.Autologous, unirradiated fibroblasts were shown in a rat model by Ferguson et al. to significantly improve healing of the irradiated surgical wound. This was evidenced by injections of culture medium, irradiated fibroblasts and non-irradiated fibroblasts in previously irradiated surgical wounds. The autologous non-irradiated fibroblasts caused greater increases of breaking load and ultimate tensile strength of the wounds than the irradiated control cells. Other aDermis-derived multipotent stem cells seem to be easily harvested from animal skin.Chunmeng reports that both topical and systemic transplantation of dermis-derived multipotent cells accelerates healing after a radiation-induced injury. The sugHuman mesenchymal stem cells have also been discussed in the therapy of radiogenic ulcers. Adipose-derived stem cells have been described for the therapy of limited local injuries and seem to improve angiogenesis and the reconstitution of dermal architecture.Akita et al. describe the use of adipose-derived stem cells (ADSC) in a female suffering from a radiogenic wound 40 years after irradiation due to a uterine carcinoma. They used an artificial dermis impregnated with ADSC to cover the wound bed and injected ADSC into the margins. This method was combined with the local administration of a basic fibroblast growth factor to improve angiogenesis. The authors documented excellent results of wound healing and a durable regenerated tissue after 1.5 years. The resA proangiogenetic effect of stem cell injections has been suggested by the authors, but other mechanisms may also serve to explain the findings in this study. New vessel formation and the optimizing of tissue microcirculation present a valuable effect of stem cell injections with respect to cutaneous wound healing. But the benefit on wound healing might end up providing a favorable environment for tumor recurrence after radiotherapy. High risk constellations or the application of stem cell injections in elderly people who have a diminished life time may put these patients at risk for developing tumor recurrence, which would justify further clinical trials. The clinical observations of Akita et al. are supported by another porcine study. Porcine skin seems to react similarly to human skin as far as radiogenic injuries are concerned. Also wound-healing mechanisms are similar in humans and pigs. That isThe antioxidant effects of adipose-derived stem cells are repoAnother advantage of adipose-derived stem cells over other stem cell sources is that they are easily obtained in large quantities by liposuction. Also their potency to synthesize growth factors and cytokines shows promise for the use in skin repair and regeneration,52. A ceThe studies mentioned above reveal that adipose-derived stem cells seem to interact directly and via paracrine mechanisms with the key cells of wound healing. Clinical trials analyzing the role of these cells and their combination with other wound-healing promoting factors would contribute to a better understanding of the potential value of this treatment regimen.The topical application of active agents to reduce the side effects of radiation therapy was analyzed in several cell culture and animal studies.Biologically active polyphenolic bioflavonoids have been shown in cell culture studies with keratinocytes to have positive effects on angiogenesis. The suggested mechanism is an increased expression of the vascular endothelial growth factor via the TNF (tumor necrosis factor)-alpha-pathway. AngiogeHistone deacetylase inhibitors  were shown to suppress the cutaneous radiation syndrome in rat skin. Chung et al. analyzed different histone deacetylase inhibitors with respect to acute skin reaction and dermal fibrosis. They used vaseline as a negative control and performed immunohistological analyses. The underlying mechanism seems to be the suppression of the aberrant expression of radiation-induced transforming growth factor-beta 1 and 2 and tumor necrosis factor alpha.This study presents the value of antitumor histone deacetylase inhibitors which may suppress cutaneous radiation syndrome and are possible new agents for increasing therapeutic gain in cancer radiotherapy.Ascorbic acid has been shown to result in the significant acceleration of healing of radiogenic ulcers. This was shown in a mouse model with doses of 10,16 and 20 Gy. A full thickness skin wound was created after radiation. Histological evaluations were performed at various times after wounding. Pretreatment with ascorbic acid augmented the synthesis of collagen significantly as revealed by an increase in hydroxyproline content. In the ascorbic acid group, an earlier wound closure was observed compared to the control group. OptimizCopper tripeptide accelerates the growth of normal and irradiated fibroblasts, which was shown in a cell culture study by Pollard et al.:Primary human dermal fibroblasts were explanted from intraoperative specimens obtained from patients after radiation therapy for head and neck cancer. Normal unirradiated fibroblasts served as a control. Irradiated fibroblasts treated with copper tripeptide showed a similar proliferation rate as the untreated controls, and produced significantly more basic fibroblast growth factor and vascular endothelial growth factor than untreated controls. An early increase in growth factors and cell proliferation by irradiated fibroblasts treated with copper tripeptide may improve wound healing.Thrombin receptor-activating peptide (P517-30) has been shown to increase wound-breaking strength in irradiated tissues. This substance is a synthesized high-affinity thrombin receptor binding peptide. In a rat model, Cromack et al. revealed that P517-30 directly stimulates resident endothelial cells and fibroblasts to overcome dermal and circulating monocytic deficits. These results suggest a mechanism to accelerate wound healing with a potential clinical application and emphasize the activity of thrombin as a growth factor. SpecialDifferent growth factors have been analyzed for their potential role in wound healing, albeit their carcinogenic potential must be taken into consideration.Important factors that may be suitable for this therapy include recombinant human granulocyte colony-stimulating factor (rhG-CSF), recombinant human macrophage colony-stimulating factor (rhM-CSF), basic fibroblast growth factor, TGF-beta and an inhibitor of transforming growth factor (TGF)-beta(1) receptor kinase-65.The exogenous application of TGF-beta (1) and (3) has the potential to act as a radioprotective agent, especially in adjuvant therapeutic regimens. In a rat model, reduced TGF-beta(3) expression was observed in the irradiated graft bed and induction of collagen synthesis was observed after application of TGF-beta (1),67.Whether these findings are relevant in the clinical setting after radiation therapy remains to be seen, because TGF-beta suppression seems to have favorable effects as well:TGF-β is a strong stimulator of extracellular matrix deposition. A new pharmacological approach was the development of the TGF-beta antisense oligonucleotide technology. This intervention shows promise as a therapeutic option for the inhibition of proteolytic tissue destruction, which is one key approach to optimize wound healing. Irradiated wounds often fail to heal because of high amounts of MMPs. TGF-beta antisense oligonucleotides seem to affect the major cell types of dermal wound healing: Fibroblasts, keratinocytes and endothelial cells were influenced in their gene expression of MMPs. MMP-1 and MMP-9 were significantly decreased after the treatment of fibroblasts and keratinocytes with TGF-beta antisense oligonucleotides in vitro. Animal TGF-beta antisense oligonucleotide technology shows promise. VEGF up-regulation was observed in vitro and a pro-angiogenetic effect of TGF-beta antisense oligonucleotides in radiation-induced dermal wounds was suggested. AnotherPlatelet-derived growth factor (PDGF) may be useful as a topical agent in post-irradiation surgical incisions,72. PDFGGranulocyte macrophage-colony stimulating factor (GM-CSF) has been shown to modulate lipid peroxidation and glutathione content in skin wounds.It has been demonstrated that GM-CSF increases the number of neutrophils, eosinophils and monocytes with corresponding bone marrow changes and might be suitable for different approaches in cancer therapy. IrradiaThe use of recombinant human epidermal growth factor (rhEGF) was studied in mice after radiation with 45 Gy. Histological examinations showed that treatment with EGF accelerated normal wound healing when compared to no treatment. Collagen distribution was significantly increased in the group treated with EGF. Dermal and epidermal structure was also more stable in the treatment group.Pollard et al. investigated cytokine expression in a cell culture model of irradiated human skin isolates. Basic fibroblast growth factor and vascular endothelial growth factor were found to be less expressed in the radiation-induced dermal wounds. These aThere is some evidence to support the combination of different growth factors to escalate their therapeutic effect. ImportaSugiyama et al. studied combined treatment in rats after local irradiation with 30 Gy. Recombinant human granulocyte colony-stimulating factor (rhG-CSF), recombinant human macrophage colony-stimulating factor (rhM-CSF) and an inhibitor of transforming growth factor (TGF)-beta1 receptor kinase, were injected into a full-thickness incisional wound site in the dorsal skin. Following combined treatment with the above three compounds the breaking strength of the irradiated skin increased to approximately one-half of that in the non-irradiated skin. Histological analysis of the wounded skin revealed an increase in the formation of collagen fibers. Moreover, the increased breaking strength was associated with an increase in a subpopulation of fibrocytes.Radiotherapy is an integral modality of head and neck cancer therapy. Compromised wound healing is an important side effect of radiation therapy. The current body of literature comprises a large number of studies investigating the molecular, cellular and clinical effects of compromised wound healing as well as current and possible future therapeutic strategies. Many of the mechanisms leading to cell injuries are still not completely understood. A better understanding of tolerance doses, improved timing of the radiation regimen, and radiation sources would allow a more focused tumor treatment. These advances have led to an improved, but not yet complete protection of healthy tissues. The clinical challenge to optimize wound healing in irradiated patients remains. The present paper critically reviews and summarizes the literature concerning the biology and possibly therapeutic strategies of radiation-induced compromise in wound healing, including stem cell injections and application of growth factors.The authors declare that they have no competing interests.FH and HG had the idea for the literature review and drafted the manuscript. EO prepared figures and tables. FP supported the manuscript by his radio-oncological knowledge. JS and HG contributed through supervision and administrative support. All authors read and approved the final manuscript.


Pungitius pungitius). In addition, we assessed the patterns of allelic differentiation between X and Y chromosomes across the populations to examine if the sex chromosomes had been highly differentiated prior to the postglacial recolonization of Fennoscandia.Sex-specific DNA markers can serve as tools for molecular sex identification, as well as for population genetic inferences. We investigated the potential utility of a microsatellite marker located on sex chromosomes for molecular sexing of Fennoscandian nine-spined sticklebacks (A clear and consistent sex difference in allele size distribution was observed at the Stn19 locus throughout the 15 populations investigated. Males were distinguishable by the presence of distinct male-specific alleles, which were lacking in all females. There was no indication of recombination between sex and the Stn19 locus in the 647 individuals tested. The degree of genetic differentiation between the X and Y chromosomes was much higher than that of interpopulation differentiation in the respective chromosomes.Our results indicate that the Stn19 locus can be used for molecular sex identification in Fennoscandian nine-spined sticklebacks. The consistent pattern of high allelic differentiation between the X and Y chromosomes in these populations suggests that the sex chromosomes were already highly differentiated prior to the postglacial recolonization of Fennoscandia. Unlike mammalian or avian species, fish exhibit diverse sex-determination systems . Even thSex-linked DNA markers can also provide significant insights into demographic history and phylogenetic relationships among populations. In particular, polymorphic Y chromosome markers are useful in tracing paternal lineages in wild populations ,15. In hPungitius pungitius) is a cold-water adapted species having a circumpolar distribution in the Northern Hemisphere and is found in a wide variety of habitats [Sticklebacks (Gasterosteidae) are widely used as model organisms in evolutionary biology -21. The habitats . This sphabitats ,23. Micrhabitats ,11,24.We investigated the potential utility of a microsatellite marker located on sex chromosomes for molecular sex identification in a large number of Fennoscandian nine-spined stickleback populations. These populations are highly subdivided from each other as a result of their postglacial colonizations following the last glacial maximum (ca. 12 000 years ago) . We therA total of 647 adult fish were collected with seine nets or minnow traps from 15 locations, comprising three coastal, two lake and 10 pond sites in Fennoscandia  individuals showed a mismatch between phenotypic and genotypic (see below) sex. All the mismatches corresponded to individuals that had been sexed on the basis of visual criteria; re-examination of these individuals by gonadal inspection confirmed their true sex, which matched the genotypic sex perfectly.FIS values were nonsignificant for females in all populations . X chromosomal alleles were polymorphic in four out of seven populations from southern and central areas (60°12'-64°34' N), but these were monomorphic in most populations from northern areas .The (δμ)s Figure . The , whereas females were characterized by the absence of these alleles. Given the male heterogamy in this species , these mDespite our large sample size, we did not find any indication of recombination between the sex-determination locus and the Stn19 locus. Consistent patterns of sex-linked allelic differentiation were observed for other microsatellite loci located on the sex chromosomes independent of their physical location . Therefore, distinct allelic divergence at the Stn19 locus between the sex chromosomes could be due to suppression of recombination between these chromosomes rather than due to tight physical linkage between the Stn19 locus and the sex-determination locus. Indeed, the sex chromosomes of nine-spined sticklebacks are cytogenetically distinguishable . Thus, bSince northern Europe was covered by the continental ice sheet until approximately 12 000 years ago, nine-spined sticklebacks must have colonized Fennoscandia after the last glacial maximum. Based on an analysis of autosomal microsatellite markers, Fennoscandian populations are highly differentiated from each other perhaps due to founder effects following postglacial colonization . Our stuThe usefulness of homologous microsatellite markers in X and Y chromosomes for the inferences of population history and sex-specific genetic structuring has been proposed in human studies ,42-44. OIn summary, we have demonstrated that the microsatellite locus Stn19 shows sex-specific allelic differences in a large number of Fennoscandian nine-spined stickleback populations. Thus, this marker can be used for the sex identification of immature individuals, as well as adults when visual criteria are deemed to be unreliable. The consistent patterns of allelic differentiation between these chromosomes across the populations suggest that the sex chromosomes of nine-spined sticklebacks had been highly differentiated prior to the postglacial recolonization of Fennoscandia.The authors declare that they have no competing interests.TS conceived the study, collected the molecular data, conducted the analyses and wrote the manuscript. GH collected the fish samples and recorded the phenotypic sex data. JM contributed to conceiving and writing of the manuscript. All authors read and approved the final manuscript.2 distances of X and Y chromosomal alleles within and between 13 nine-spined stickleback populationsMatrix of (δμ).Click here for file


Type 2 diabetes mellitus is a metabolic noncommunicable disease with an expanding pandemic magnitude. Diabetes predisposes to lower extremities ulceration and impairs the healing process leading to wound chronification. Diabetes also dismantles innate immunity favoring wound infection. Amputation is therefore acknowledged as one of the disease's complications. Hyperglycemia is the proximal detonator of systemic and local toxic effectors including proinflammation, acute-phase proteins elevation, and spillover of reactive oxygen and nitrogen species. Insulin axis deficiency weakens wounds' anabolism and predisposes to inflammation. The systemic accumulation of advanced glycation end-products irreversibly impairs the entire physiology from cells-to-organs. These factors in concert hamper fibroblasts and endothelial cells proliferation, migration, homing, secretion, and organization of a productive granulation tissue. Diabetic wound bed may turn chronically inflammed, procatabolic, and an additional source of circulating pro-inflammatory cytokines, establishing a self-perpetuating loop. Diabetic fibroblasts and endothelial cells may bear mitochondrial damages becoming prone to apoptosis, which impairs granulation tissue cellularity and perfusion. Endothelial progenitor cells recruitment and tubulogenesis are also impaired. Failure of wound reepithelialization remains a clinical challenge while it appears to be biologically multifactorial. Ulcer prevention by primary care surveillance, education, and attention programs is of outmost importance to reduce worldwide amputation figures. The former is a condition in which by autoimmune mechanisms pancreatic ficiency . T2DM usficiency .Surgeon Davies Pryce put forward as early as 1887 the link between diabetes and foot ulceration by writing in The Lancet that “diabetes itself may play an active part in the causation of perforating ulcers” . HoweverκB), thus amplifying the absurd inflammation and intoxicating the wound milieu [http://www.bioline.org.br/) data sources.The healing process in diabetes is also jeopardized by the patient's susceptibility to infection due to deficiencies on the innate immunity. Although the diabetic wound bed may be adversely overwhelmed by inflammatory cells, it does not represent an overt antibacterial protection. By the contrary, the diversion of glucose to the polyol pathway affects bacterial killing by reducing neutrophil opsonophagocytosis. Furthermore, hyperglycemia-induced reactive oxygen species (ROS) deregulated the innate immunity via an overactivation of NF-kappaB . They also secrete growth factors as important messengers for mesenchymal-to-mesenchymal and epithelial-mesenchymal communication, especially for establishing the emerging basement membrane and subsequent reepithelialization. Therefore, any impediment to fibroblast function is deterrent for normal wound healing and may result in chronic, nonhealing wounds. The fibroblast, when engaged in fibrogenesis, displays the highly activated phenotype characteristic of myofibroblasts. Although their origin has not yet been definitely elucidated; proliferation of preexisting adjacent dermal fibroblasts and, probably, recruited from the bone marrow has been documented . Under tbroblast .in vitro models that demonstrated that in diabetics' cutaneous fibroblasts; synthetic, proliferative, and secreting capabilities are reduced [in vitro, but that the aging process appeared more precociously than in nondiabetic counterparts [in vitro, full-thickness wounds induced in nondiabetic pigs exposed to a local hyperglycemic environment exhibited no difference in wound closure when compared with normoglycemic controls, suggesting that delayed wound healing by diabetes is a far more complex phenomenon than circumscribed high-glucose concentration itself [Rowe and coworkers pioneered the  reduced . Other p reduced . Followi reduced . Years lterparts . Other sterparts . This reterparts , thus coterparts . Accorditerparts . In additerparts . Most ofterparts . In contn itself . As a con itself . Using tn itself . Enrichiα1β1) and not through the canonical receptor for advanced glycation end-products (RAGE) [Oxidative stress appeared dependent on the upregulation of NAD(P)H oxidase 4 (Nox4), a reactive oxygen species Nox homologue, which appeared activated by p38 MAPK. Proximal to this cascade is the effect caused by the interaction of the modified collagen with 3DG, which signals to the fibroblast by interacting with integrins alpha-1/beta-1 (s (RAGE) . Other gs (RAGE) . A subses (RAGE) . Under aIn addition to the deleterious effects of glucose and its derivatives, diabetic fibroblasts exhibit particular features. Literature documents that diabetic mice fibroblasts show a severe impairment in VEGF production under normoxic and hypoxic conditions in addition to an increased pro-degradative activity due to the high expression of matrix metalloprotease type 9 (MMP-9) . Similarα-) induced apoptosis both of which are abundant in diabetic connective tissue [α/SDF-1α) observed in experimental and clinical wounds when insulin is topically administered may be attributable to FOXO1 neutralization. Curiously, the acceleration of wound healing occurs in parallel to a local recovery in the expression of proteins involved in insulin signaling pathways [It seems that amplification of oxidative stress acts as a primary culprit in harming fibroblasts biology in diabetes, involving electron transport in mitochondria. High intracellular glucose levels increase the electron transport chain in mitochondria during oxidative respiration, leading to formation of O2- and the generation of various reactive oxygen species derivatives in the mitochondria. Other sources of oxidative stress in diabetes include glucose autooxidation, the polyol pathway with ensued depletion of antioxidant reserves and the formation of AGEs . Chronice tissue . FOXO1 le tissue , 36. Intpathways . Aside fpathways , which eex vivo diabetics' fibroblasts behavior.Why diabetics' fibroblasts evoke behavioral traits in culture mirroring the donor's tissue, even when grown under optimized oxygenation, nutrient, growth factors, and glucose supply?Is there any sort of “behavioral imprinting” so that they are reminiscent from a diabetic donor?Why cultured fibroblasts from both ischemic and neuropathic ulcers exhibit different ultrastructural morphology and organize the monolayer in a privative manner? in vivo traits?Is there any epiphenomenon beyond the irreversible glycation sustaining the “impersonation” of the Despite the prolific investigation conducted during all these years, still questions remain to be answered in relation to β (TGF-β), in collaboration with other proteins as integrins, cadherins, and ephrins, regulate angiogenesis by promoting endothelial cells recruitment, proliferation, migration, coopting, and collar stabilization. There is an enormous and ever-growing body of evidence indicating the close correlation between hyperglycemia and the abnormalities in endothelial function and morphology [Angiogenesis is a comprehensive term that indicates the physiological process involving the growth of new blood vessels or neovascularization. This is a vital process for embryological growth, tissue development, and wound healing. Different growth factors families as vascular endothelial growth factors (VEGF), fibroblast growth factor (FGF), angiopoietins, platelet-derived growth factor (PDGF), transforming growth factor-rphology . The UK rphology . For subrphology . Furtherrphology .per se seem to be a very sensitive target to high glucose. Endothelial dysfunction is intricately related to insulin resistance through the stimulatory effects of insulin on glucose disposal and NO production in the endothelium. Today, vascular dysfunction remains as a major cause of morbidity, amputation/disability, and mortality in diabetic patients. Even after achieving the successful reperfusion of an ulcerated lower extremity, the healing process is slow and torpid. Therapeutic angiogenesis has been pursued for years, but the clinical results have shown relatively limited outcomes [in vitro and in vivo and as for multiple physiological processes; insulin and its downstream signaling regulate most of the endothelial cell functions [In vitro models simulating “normoglycemia” and “hyperglycemia” have demonstrated that under high glucose ambient, proliferation and tube formation of dermal microvascular endothelial cells appear impaired [α level concomitant to the death receptors TNF-R1 and Fas in a variety of cultured endothelial cells [As depicted for fibroblasts, high glucose and the glycated by-products exert a toxic effect on endothelial cells and the vascular wall in general. In parallel, the endothelial cells outcomes –45. Highunctions . High glunctions . In addiunctions  thus conimpaired . Furtherimpaired . High glal cells . Under tal cells .α, interleukin-1β (IL-1β), and interleukin-6 (IL-6), ultimately leading to decreased insulin sensitivity and diabetic vascular complications. Moreover, hyperglycemia-induced FOXO also plays an important role in the induction and amplification of proinflammatory cytokines production. FOXO1 directly binds to IL-1β promoter and increases its expression in macrophages [Hyperglycemia-induced oxidative stress promotes inflammation through increased endothelial cells damage, microvascular permeability, and uncontrolled release of proinflammatory cytokines, including TNF-rophages .β [Hyperglycemia and the accumulation of AGEs disturb the role of angiogenic growth factors as VEGF, and its receptor, its signaling pathway, thus disrupting endothelial proliferation, migration, and endothelial progenitor cells (EPCs) recruitment and release from bone marrow . Insulinβ . Studiesβ .in vivo. The fact that angiogenesis is restored by aminoguanidine treatment reinforces the antiangiogenic role of AGEs [The glycation of collagen and other proteins within the wound extracellular matrix and AGEs accumulation bring catastrophic consequences for the angiogenic reaction with inhibition of angiogenesis  of AGEs . Angioge of AGEs . This evin vivo models [κB and consequently the expression of the inducible form of nitric oxide synthase (iNOS), intercellular adhesion molecule-1 (ICAM-1), and other inflammatory mediators [Compelling evidence indicates that at least a portion of the hyperglycemia-mediated endothelial damages and dysfunctions are associated with an impaired mitochondrial activity resulting in mutations of mitochondrial DNA, due to a disproportionate reactive oxygen radicals production, leading to an inflammatory reaction and apoptosis . As a mao models . Many ofo models . Hyperglo models . The incediators .in vitro as to organize tubulogenic structures when subcutaneously implanted in a matrigel plug, thus hindering the revascularization of damaged areas [in vivo by limiting cells proliferation and differentiation [κB, and caspase-3 in a MAPK- (ERK/P38/JNK-) dependent manner [Endothelial Progenitor Cells (EPCs) are active players for the maintenance and repair of endothelial cells. They participate in angiogenesis as they proliferate, migrate, and differentiate and are a source for proangiogenic factors and cytokines . Multipled areas . Over agntiation . As to fntiation . As desct manner .how and why microvascular morphological changes that recreate chronic, life-time processes are readily identified in a 7-day-old granulation tissue fragment, even in compensated patients. This incites to investigate which are the diabetes' operational local and/or systemic forces that can disrupt vascular morphogenesis.The diabetes-mediated vascular damage is perhaps the most outspoken and ancestrally identified emblem of diabetes. It is varied and broad as it is the concept of systemic endothelial dysfunction. Diabetes distorts the angiogenic program to ironically culminate with a maldistribution of soluble angiogenic factors: shortage where and when required (lower extremities skin) but overproduced where and when not needed (retina). It is also challenging, to understand Once we have described the main consequences of high glucose/hyperglycemia on the two principal architects of the granulation tissue: fibroblasts and endothelial cells, we are intended to recapitulate the most distinguishing features on the onset of the granulation process in diabetic cutaneous wound healing.Tissues' regenerative capabilities have been neglected along the species evolution; thus, scarring process has emerged as an urgent alternative to favor the structural and functional restoration of a wounded zone. Within these events, the process of granulation tissue formation is pivotal as it constitutes a sort of living, temporary aggregate of cells and proteins, acting as a welding material until the tissue's continuity is restored. However, the reluctance to trigger and sustain the out-growth of a productive granulation tissue with an appropriate extracellular matrix is typical in diabetic patients, and particularly if ischemia concurs. As mentioned, these wounds are characterized by a proliferative arrest, proinflammed, prooxidant, and prodegradative phenotype .per se is a metabolic disease in which fuels metabolism is perturbed given the rupture of one of the most important anabolic axis of the organism: insulin/insulin-like growth factor type-I. The role of insulin in wound healing is well known by its anabolic effect on wound protein balance favoring synthesis and preventing degradation [This stubbornness and slowness to heal in diabetes is conditioned by systemic and local factors that in complicity counteract intrinsic reparative mechanisms. In a broad systemic context, inflammation and the anabolic deficit can be conceptually mentioned. Diabetic patients with foot ulceration bear a specific and nonrandom alteration of the immune status with an active upregulation of circulating levels of acute-phase proteins, cytokines, and chemokines that impose a chronic systemic inflammatory profile and amplify local wound inflammatory networks . The sysradation , 75. IGFradation . We do nradation . In suppradation .α, rapidly undergoing premature senesce, arrest or apoptosis. Fibroblasts are the main source of collagen, and the number of fibroblasts can be taken for a measure of repair by their collagen synthesis ability. It is very likely that the deficit of growth factors such as TGF-β1, IGF-I and PDGF could translate into a DFU with scarce extracellular matrix acumulation and impoverished cellularity. Numerous growth factors  are able to regulate the balanced expression of matrix metalloproteases and tissue inhibitors of metalloproteases (MMPs/TIMPs), while most of them exhibit an altered expression in DFU [β1 and TGF-β3, in which the former appears downregulated, may explain fibroblasts quiescence in terms of proliferation and secretion [Rapid formation and deposition of an appropriate extracellular matrix, in particular by fibroblasts, is required for an efficient cellular anchoring and homing at the wound bed. As mentioned above, the cutaneous fibroblast is a sensitive cell to high glucose, AGE-precursors, AGEs, ROS, and TNF-n in DFU . Moreoveecretion . This phecretion . The extecretion . One of ecretion . We deemecretion  may be iThe diabetic granulation process does not generally exhibit the orderly cascade of events that characterize normal wound healing. This has been confirmed through the histopathological analysis of granulation tissue biopsies by Loots and coworkers who described the lesions as “frozen” in a chronic low-grade inflammatory state associated to a scarce provisional extracellular matrix . Our groα and IL-1β, which act as a triggering signal for MMPs expression via the common NF-κB signaling pathway. Within the wound context, TNF-α stimulates its own secretion and that of IL-1β, which contributes to a persistent inflammatory status [α has proved to negatively impact the repair process as it is early secreted since the inflammatory phase. Its deregulation is not only associated with persistent inflammation but also to connective tissue degradation [α mediates its antagonistic effects on TGF-β1 through the JNK pathway via inhibition of Smad phosphorylation, consequently reducing the expression of TGF-β1, and that of several downstream matrix proteins [Under more mature stages, the neuropathics may also show an abnormal sprout of new small vessels and capillaries that may derive not from a normal angiogenic response but due to arteriovenous shunts. Our observations remind us with those of Black and coworkers who demonstrated that in neuropathic patients there exists a decrease in fibroblast proliferation and a scarce amount of collagen accumulation within the wound bed . On the y status . TNF-α hradation . Concomiproteins . In thisproteins .α5β1 to effectively migrate [Importantly, the perpetuated homing of PMN within the wound bed is associated to high local levels of elastase secretion, ROS, and reactive nitrogen species . High ci migrate . Curious migrate . The con migrate .α stands as a major driving force. The negative impact of TNF-α level on the sensitivity of tissues to insulin has been consistently documented. Skin cells are not excluded from this effect [α or to increase the wound local availability of active TGF-β1 would be similarly effective for stimulating granulation tissue and wound closure [The link between wound cells and apoptosis was described above; we just wish to comment that in sharp contrast to wound-infiltrated inflammatory cells, which become refractory to apoptosis, granulation tissue-producing cells are sensitive to commit suicide where TNF-s effect . Conclus closure .Chronic wounds and especially diabetic foot ulcers exhibit a highly pro-oxidant microenvironment that amplifies the cytotoxic cascade. Endothelial cells and fibroblasts, in particular senescent fibroblasts, are a prominent source for oxygen radicals, but at the same time they turn into these radicals targets which by converging mechanisms arrest cell proliferation and induce apoptosis . Thus, tα-SMA). Nevertheless, α-SMA expression is precisely controlled by the joint action of growth factors like TGF-β1 and extracellular matrix proteins like the fibronectin (FN) splice variant ED-A, as by the local mechanical microenvironment [β1, fibronectin, and other matrix proteins may appear in deficit. Hence, all these factors may contribute to the poor contractile activity. Furthermore, Goldberg and coworkers have shown that among the deleterious activities of TNF-α within the wound is to suppress α-SMA expression in human dermal fibroblasts [The failure of wound contraction is a clinical hallmark of diabetic granulation tissue. Fibroblast-to-myofibroblast transdifferentiation represents a key event during wound healing and tissue repair. The contractile force, generated by myofibroblasts as a highly specialized cell, speeds the healing process of dermal wounds in healthy humans, accounting for an 80–90% of scar tissue reduction . In addiironment . It shouroblasts .  Figureα), which acts as a recruiting factor and its CXCR4 chemokine receptor are also impaired by diabetes [If the animals-derived evidence that a high fraction of the wound myofibroblasts potentially derives from bone marrow fibrocytes is valid for humans ; we havediabetes . Finallydiabetes .What are the molecular and cellular driving forces supporting the microscopic structural differences between neuropathic and ischemic ulcers beds?What is the explanation for the “inheritance” of vascular changes as a dramatic Monckeberg media thickening in nascent arteries within an early hatching granulation tissue?Why granulation tissue is histomorphologically abnormal even in metabolically compensated patients?Reepithelialization at the clinical level, it is not a lesser important problem as most of the diabetic wounds may granulate in time, while reepithelialization is even far slower, arrhythmic, and torpid. Reepithelialization is accomplished through the combined actions of keratinocytes' dedifferentiation, proliferation, and migration requiring a complex basement membrane, emerged of the mutual interaction between mesenchymal and epithelial cells. Reepithelialization failure is therefore one of the landmarks of diabetic and other chronic wounds. The epidermal edge of a chronic wound is thick and hyperproliferative with mitotically active keratinocytes unable to migrate along the surface, and by the contrary, moving down deep into the neodermis. Therefore, it has been speculated that the nonhealing edge keratinocytes do not successfully complete either of two possible pathways: activation or differentiation. In consonance with this, one of the major issues in chronic wounds treatments is how to revert the chronic wound keratinocytes' phenotype to a proper differentiating and migratory program [There are numerous cellular and molecular aspects unknown and that remain to be answered on the granulation tissue biology. program .Glucose has shown to exhibit a direct toxic effect on keratinocytes. As for other cells grown in the presence of high glucose concentrations, human epidermal keratinocytes significantly reduce their proliferation rate and replicative life span and were rendered more susceptible to commit apoptosis . Other skt1-mediated posttranscriptional mechanism [in vivo [α has also been implicated in epithelial cells arrest by deeply perturbing critical elements of keratinocytes' physiology, including insulin sensitivity [The fact that insulin is biologically relevant for skin cells derives from the observation that insulin is an essential component for cultured human keratinocytes, demonstrating its involvement in the regulation of proliferation, survival, and metabolism . Recent echanism . Glucoseechanism . Lack of[in vivo . Furthersitivity .β-catenin in impairing epithelial edges migration. Nuclear beta-catenin stabilization inhibits keratinocytes migration by blocking epidermal growth factor response via c-myc induction, and repressing keratins 6 and 16 expression, depleting at the end the pool of epidermal stem cells at the nonhealing edge [in vitro and in vivo [γ-1 (PLC-γ1). On the other hand, EGFR activation also leads to membrane ruffling and focal adhesions through activation of members of the Rho subfamily of GTP-binding proteins [A notorious study has provided evidence aiming at the roles of c-myc and ing edge . It is ting edge . Moreove in vivo . Among t in vivo , with thproteins . Recent proteins . Classicproteins  for reepproteins . As diabproteins . At the Above all, questions from the clinical practice remain and pose as a challenge for basic researchers: (1) why after wound contours surgical debridement keratinocytes migration resumes, for soon after become stunted and arrested again? (2) Why the biopsies invariably show a hypertrophic lip of epithelial cells in vertical, downward growth in spite of a horizontal polarization?per se is a complex disease, our contemporary understanding on the molecular mechanisms impairing wound healing in diabetes has indefectibly expanded over the last 20 years. The last few years have witnessed the birth of the notion that type 2 diabetes is not a single, unique process, whereas the concept of group of diseases has flourishing. Type 2 diabetes seems to be ethiopathogenically multifactorial and behaves as individual as the affected subject is. So is the pattern of the clinical complications, including the wound itself.Although diabetes To date, all the evidence aims to high glucose burden as the proximal trigger to unleash acute and chronic self-perpetuating loops, which include but are not limited to ROS-lipid peroxidation, hyperinflammation/disimmunity, AGE-RAGE toxicity, mitochondrial dysfunction, and nytrosilation end-products accumulation. The concerted action of these factors enforces fibroblasts, pericytes, and endothelial cells to a precocious senescence, arrest, and apoptosis. Indeed, the failure of the agonistic stimulation of the insulin axis deeply impacts on the biology of diabetics' cells. This is a seminal axis that connects the anabolic role of insulin via aminoacids transporters translocation and protein synthesis with cell survival and proliferation mechanisms, thus preventing apoptosis, autophagy (cells-self catabolism), and arrest. Fatal for the cells is silencing the agonistic stimulation of tyrosine kinase-growth factors receptors which would ensure cell cycle progression. At both experimental and clinical levels, the diabetic wound phenotype is the expression of countless molecular factors that operate through a complex biochemistry and lead to an aberrant cellular behavior. The pathway to chronification has not been fully elucidated but by all means it represents a form of cells' biological disobedience and entails the need of continuous surgical “cuttings” in order to transiently restore an acute behavior by “refreshing” the cellular environment. It is likely that the scarcity of insulin and growth factors-induced tyrosine kinase receptors downstream signaling may lie, at least in part, behind chronification. Diabetes takes away the resources that ensure wound cells perpetuation and turn-over.Although type 2 diabetes worldwide expansion is undeniable, primary care ulcer prevention plans together with the emergence of first-line pharmaceuticals and smart devices like engineered skin equivalents will certainly prevent and reduce contemporary amputation figures.


Clinical decision for primary treatment for prostate cancer is dictated by variables with insufficient specificity. Early detection of prostate cancer likely to develop rapid recurrence could support neo-adjuvant therapeutics and adjuvant options prior to frank biochemical recurrence. This study compared markers in serum and urine of patients with rapidly recurrent prostate cancer to recurrence-free patients after radical prostatectomy. Based on previous identification of urinary sarcosine as a metastatic marker, we tested whether methionine metabolites in urine and serum could serve as pre-surgical markers for aggressive disease.Urine and serum samples , collected at the time of prostatectomy were divided into subjects who developed biochemical recurrence within 2 years and those who remained recurrence-free after 5 years. Multiple methionine metabolites were measured in urine and serum by GC-MS. The role of serum metabolites and clinical variables  on biochemical recurrence prediction were evaluated. Urinary sarcosine and cysteine levels were significantly higher (p = 0.03 and p = 0.007 respectively) in the recurrent group. However, in serum, concentrations of homocysteine (p = 0.003), cystathionine (p = 0.007) and cysteine (p<0.001) were more abundant in the recurrent population. The inclusion of serum cysteine to a model with PSA and biopsy Gleason grade improved prediction over the clinical variables alone (p<0.001).Higher serum homocysteine, cystathionine, and cysteine concentrations independently predicted risk of early biochemical recurrence and aggressiveness of disease in a nested case control study. The methionine metabolites further supplemented known clinical variables to provide superior sensitivity and specificity in multivariable prediction models for rapid biochemical recurrence following prostatectomy. Prostate cancer remains the most common non-cutaneous solid malignancy in the United States, and the second leading cause of cancer specific death in men. Nevertheless, it has become increasingly clear that not all men who are diagnosed with prostate cancer require intervention A previous study identified sarcosine (N-methylglycine) as a product of methionine catabolism that is elevated in the urine of patients with metastatic prostate disease In this study we evaluated the serum and urine of radical prostatectomy patients for metabolites to differentiate those who developed early biochemical recurrence (rise in serum PSA≥0.2 ng/ml) within two years of surgery and those who remained recurrence-free after more than five years. We found that the urine of patients in the rapidly recurrent group had significantly higher concentrations of sarcosine and cysteine than those in the recurrence-free group. In addition, significantly greater concentrations of serum cystathionine, homocysteine and cysteine were found in the rapidly recurrence group compared to the recurrence-free group. These products of elevated methionine catabolism in patients with rapidly recurrent prostate cancer represent pre-surgical indicators that augmented serum PSA for the prediction of clinically significant prostate cancer.This nested case-control study was conducted in accordance with the Institutional Review Board of Vanderbilt University. Written consent was given by the patients for their information to be stored in the hospital database. The board specifically approved the research use of the di-identified information and “on the shelf” specimens to be used for research under a waiver of consent.The digital medical records of 400 subjects were retrospectively examined using the Vanderbilt University Department of Urologic Surgery registry of radical prostatectomies performed between 2003 and 2007. Several patients were excluded for reasons of compromised renal, heart, or liver function as was determined by electronic records of elevated urinary creatinine, hypertension, cardiac infarction history, and blood markers for hepatic function. Additionally, availability of follow-up data and records of pre-surgical hormone-ablation therapy were reasons for exclusion. Rapidly recurrent patients were identified as those who developed biochemical recurrence following prostatectomy within 2 years . The recurrence-free population was defined as having maintained a serum PSA<0.01 ng/ml for five or more years following surgery. Ultimately, for this nested case control study we focused on 54 subjects for analysis of urine and 58 subjects for analysis of serum who developed rapid biochemical recurrence and an age-matched recurrence-free control group who were free of recurrence. The mean age for the subjects was 60 years. All subjects were annotated based on age, pre-surgical serum PSA, biopsy Gleason score, clinical stage, and detection of biochemical recurrence.Serum and urine obtained at the time of radical prostatectomy were rapidly processed and stored at −80°C. We evaluated serum and urine for the metabolites, sarcosine, dimethylglycine, methionine, homocysteine, cystathionine, cysteine, methylmalonic acid and methylcitrate by gas-liquid chromatography/mass spectrometry Patients' baseline demographic and clinical variables were assessed using Wilcoxon rank sum tests for continuous variables and Fisher exact tests for categorical (including binary) variables. All marker values, as well as PSA levels, were logarithmically transformed to achieve normality. Correlations among the markers were assessed using Spearman's rank correlation. Logistic regression models were used to analyze incidence of recurrence. The base model includes serum PSA, biopsy Gleason score, and clinical stage, clinical variables that are available prior to surgery. The post-surgical variables  were not considered. For multiplicity control, p≤0.007  was considered statistically significant. To avoid further overfitting of the data, no variable selection was performed in the subsequent analyses based on logistic regression models. We used a likelihood ratio test to compare the simpler model (without the metabolites) and the full model . Receiver operating characteristics (ROC) curves were generated for each logistic regression model, where the area under ROC curve (AUC) was determined. Integrated discrimination improvement (IDI) and Net reclassification index (NRI) Urine metabolites were initially measured in fifty-four patients who developed biochemical recurrence (N = 25) and those that remained recurrence-free (N = 29). These patients were matched for age and pre-surgical serum PSA. We then performed a nested case control study with pre-surgical serum. Fifty-eight age-matched prostatectomy patients were stratified by pre-surgical PSA, clinical stage, and biopsy Gleason grade as well as pathologic variables Table 2.The relevance of these newly identified markers to patient recurrence status were illustrated in Kaplan-Meier plots for homocysteine, cystathionine, and cysteine as compared to pre-operative serum PSA levels, and time-to-recurrence Figure 12 = 0.65, p<0.01), and cysteine was also highly correlated with cystathionine . Based on multiple logistic regression models (The clinical value of these methionine metabolites as biomarkers would be to significantly increase the ability to predict aggressive prostate cancer features and early biochemical recurrence over and above existent clinical variables including serum PSA, biopsy Gleason score, and clinical stage. We developed a multiple logistic regression model for the prediction of biochemical recurrence based on serum methionine metabolites and the pre-surgical predictor variables, serum PSA and biopsy Gleason grade. Since majority of patients in both cohorts had clinical stage T1c disease, this variable had little discriminative power and was dropped from the model. Serum cysteine, cystathionine, and homocysteine were the top three predictors for recurrence in 70% of the patients, so further analysis of methionine metabolites focused on these three metabolites. Correlations between cysteine and homocysteine were the highest among all pair-wise correlations (Rn models Table 5,To define the efficacy of the markers in predicting recurrence-free survival, Cox proportional hazard regression models were fit showing that cysteine, cystathionine, and homocysteine were each independent predictors of recurrence-free survival when adjusting for pre-operative serum PSA and biopsy Gleason score Table 7.Current risk stratification of patients prior to surgery involves variables including serum PSA, clinical stage, and biopsy grade. Independent serum markers in conjunction with PSA could help distinguish patients with aggressive prostate cancer. In the current era of PSA testing, clinical staging has reduced relevance when tumor volumes are relatively small. In our study, the highest biopsy Gleason score in ≥8-core biopsies provided a significant independent predictor comparable to serum cysteine and homocysteine. However, routine ultrasound directed first biopsies are reported to miss nearly a quarter of the prostate cancers Pathways of methionine metabolism involve two mechanisms for sarcosine formation Figure 3Gnmt null animals were 33 fold higher than in the livers of wild type animals and all of the Gnmt null animals developed hepatocellular carcinoma after 8 months The majority of the sarcosine produced in the body is made in the liver as a downstream product of SAM and homocysteine. Studies using homozygous mice with GNMT knocked out have plasma SAM levels 50% greater than that of wild type. The SAM levels in the livers of the To our knowledge, no previous study has correlated an entire arm of a metabolic pathway in the aggressiveness of cancer. In our study the comparison was made between patients with proven cancer, not between subjects with proven cancer and benign prostatic disease. These results were obtained with only a relatively small group of patients but the results are significant and suggest that further studies are needed. The underlying biology supports the robustness of these markers. Higher serum homocysteine, cystathionine, and cysteine improved the utility of currently used clinical variables in predicting early recurrence and suggest a greater flux of methyl groups through the enzyme GNMT.