Biological markers in inflammatory bowel disease: practical consideration for clinicians.

The biomarkers are important in the Inflammatory Bowel Disease (IBD) to gain an objective measurement of disease activity and severity, as well as prognostic indicator and outcome of therapy. And they can be helpful to avoid invasive procedures. The ideal biomarker does not exist for IBD and it is likely that more than one biomarker will be needed. Biological markers potentially useful in IBD include acute-phase proteins, fecal markers, several antibodies and novel genetic determinants. The C-reactive protein (CRP) is the most studied and has been shown to be an objective marker of inflammation. CRP is a good marker of measuring disease activity in Crohn's disease (CD) and its levels can be used to guide therapy. The fecal markers (calprotectin and lactoferrin) may be helpful in differentiating patients with IBD from those with functional disorders and to predict clinical relapse. The panel of serologic markers (anti-Saccharomyces cerevisiae antibody, perinuclear anti-neutrophil cytoplasmic antibody, anti-OmpC and anti-I2 and antiglycan antibodies) for IBD can be used to stratify IBD patients into more homogeneous subgroups with respect to disease progression. Correlating serologic markers with genotypes and clinical phenotypes should enhance our understanding of the pathophysiology of IBD. The development of biomarkers in IBD will be very important in the future with the increasing utilization of novel methodological approaches like genomics and proteomics.

Higher infliximab trough levels are associated with perianal fistula healing in patients with Crohn's disease.

BACKGROUND: Infliximab has been found to be efficacious in the treatment of fistulas in the setting of Crohn's disease, even though some patients do not benefit from therapy. AIM: To assess the correlation between perianal fistula healing and trough levels of infliximab. METHODS: In this cross-sectional study, we identified patients with Crohn's disease who had perianal fistulas and were treated with infliximab for at least 24 weeks. We excluded patients who underwent a faecal diversion procedure or proctectomy. Predictive variables included demographics, disease phenotype, disease activity, infliximab levels, anti-infliximab antibodies. The primary outcome was fistula healing defined as the absence of drainage. The secondary outcome was complete fistula closure and mucosal healing. RESULTS: 117 patients were included. Patients with fistula healing had significantly higher median serum infliximab levels when compared to those with active fistulas [15.8 vs. 4.4 µg/mL, respectively (P < 0.0001)]. There was an incremental gain in fistula healing with higher infliximab levels. The AUC for the association between fistula healing and infliximab levels was 0.82 (P < 0.0001), while the AUC for the association of infliximab levels and fistula closure was 0.69 (P = 0.014). Patients with anti-infliximab antibodies had a lower chance of achieving fistula healing (OR: 0.04 [95%CI: 0.005-0.3], P < 0.001). CONCLUSIONS: There is a significant association between serum infliximab levels and rates of fistula healing. Achieving infliximab levels ≥10.1 mcg/mL in patients with Crohn's disease and perianal fistulas may improve outcomes as part of a treat-to-target strategy.

Inflammatory bowel disease is presenting sooner after immigration in more recent US immigrants from Cuba.

BACKGROUND: Despite a rising incidence of inflammatory bowel disease (IBD) in Hispanics in the United States, there are no studies examining the relationship between immigrant generation and IBD onset among Hispanics. AIMS: To determine whether age of IBD diagnosis, time from immigration to IBD diagnosis and IBD phenotype, differed across immigration periods in South Florida Cuban immigrants. METHODS: This was a cohort of consecutively identified Cuban-born adults who developed IBD in the United States and were followed in gastroenterology (GI) clinic. We divided time cohorts of immigration by historical relevance: before 1980, 1980-1994 and 1995-to-present. We examined differences across time cohorts in diagnosis age, time from immigration to IBD diagnosis, and IBD phenotype (ie, IBD type, disease location). RESULTS: A total of 130 Cuban patients with IBD were included. Age of IBD diagnosis was older in Cubans arriving before 1980 than in those arriving between 1980-1994 or after 1995 (44.7 vs 33.79 and 33.71, respectively, P<.0001). Time between immigration and diagnosis was shorter in patients arriving to the US after 1980 (31.77 years, Standard deviation (SD) 12.83 (<1980) vs 17.13 years, SD 8.55 (1980-1994) and 8.30 years, SD 4.72 (1995-to-present). IBD phenotype, including type of IBD, disease location and surgeries, did not differ significantly across time cohorts. CONCLUSIONS: Our study describes changing patterns of IBD onset following immigration in Cubans, suggesting that environmental changes either in the United States, Cuba or both are resulting in faster IBD onset in younger immigrant generations. These studies can inform the search for environmental triggers that may result in IBD.

Efficacy of mycophenolate mofetil in patients failing 6-mercaptopurine or azathioprine therapy for Crohn's disease.

Mycophenolate mofetil (MMF) is a novel immunomodulator that may be effective in the treatment of chronic active and perianal Crohn's disease (CD). The aim of this study is to assess the efficacy of MMF in CD patients who failed or were intolerant of 6-mercaptopurine (6-MP) or azathioprine (AZA). Eleven CD patients were treated with MMF after a failed course of 6-MP/AZA, and their records reviewed retrospectively. Reasons for 6-MP/AZA intolerance or failure were recorded. Response to MMF was determined by calculation of the Harvey-Bradshaw index and ability to taper steroids. Adverse reactions to MMF were recorded. Eleven patients were identified who failed a previous trial of 6-MP/AZA and other immunomodulators and required immunomodulator therapy. Of 11 patients who started MMF, four had early adverse reactions within 8 weeks and stopped the medication. Of the remaining seven patients who took MMF for at least 8 weeks, one had a complete response, two had a partial response, and four had no response to the medication. In patients who failed 6-MP/AZA, MMF was of benefit in 3 of 11 patients with only one complete responder. This lower-than-expected response rate may indicate that patients who are resistant to 6-MP or AZA may also be resistant to MMF.

An open-label pilot study using thioguanine as a therapeutic alternative in Crohn's disease patients resistant to 6-mercaptopurine therapy.

BACKGROUND AND AIMS: A substantial number of patients with inflammatory bowel disease (IBD) fail to achieve a complete clinical response with 6-mercaptopurine (6-MP) and azathioprine (AZA). Inability to achieve therapeutic 6-thioguanine nucleotide (6-TGN) levels due to the preferential overproduction of 6-methylmercaptopurine ribonucleotides (6-MMPR) upon dose escalation characterizes a newly described subgroup of IBD patients resistant to 6-MP/AZA therapy. Treatment with 6-thioguanine (6-TG), a related thiopurine, which forms 6-TGNs more directly may be beneficial in such patients. This pilot study evaluated the safety, tolerance, and efficacy of 6-TG in the subgroup of Crohn's disease (CD) patients failing to attain adequate disease control with traditional 6-MP/AZA therapy. METHODS: Ten CD patients with preferential 6-MMPR production upon 6-MP/AZA dose escalation were enrolled in an open-label pilot study. Seven of 10 patients had experienced dose-related 6-MP toxicities. RESULTS: Seventy percent of the patients (7 of 10) responded or were in remission at week 16. Clinical response was evident by week 4 in most. 6-TGN levels were nine-fold higher with 6-TG treatment than with 6-MP, whereas 6-MMPR levels were undetectable. No patient developed a recurrence of hepatic or hematological toxicity. CONCLUSIONS: 6-TG was a safer and more efficacious thiopurine in this subgroup of IBD patients resistant to 6-MP therapy. Larger controlled trials are warranted to further evaluate both the short- and long-term safety and efficacy in this subgroup of patients as well as a broader spectrum of IBD patients.

Infliximab in the treatment of medically refractory indeterminate colitis.

AIM: To examine the outcome of infliximab intervention in refractory indeterminate colitis. METHODS: Twenty patients with severe, medically refractory indeterminate colitis were treated with infliximab. All patients initially received infliximab, 5 mg/kg, intravenously and, in some patients, the dose was subsequently increased to 10 mg/kg. The number of infusions ranged from one to 16 per patient. Indeterminate colitis was defined as colitis that could not be classified with certainty as Crohn's disease or ulcerative colitis based on traditional clinical, endoscopic and histopathological criteria. The clinical response to infliximab was classified as complete response, partial response or non-response. RESULTS: Fourteen of the 20 patients (70%) showed a complete response to infliximab treatment, two showed a partial response and four showed no response. The four non-responders underwent colectomy with ileal pouch-anal anastomosis. The resected colon specimen was consistent with ulcerative colitis in all four cases, although two were subsequently re-classified as Crohn's disease. Eight additional patients were subsequently re-classified as having Crohn's disease on longer follow-up evaluation, whilst eight continued to have features of indeterminate colitis. The response rate to infliximab treatment was similar in both groups. CONCLUSIONS: Infliximab is effective in approximately two-thirds of patients with indeterminate colitis, and thus may be considered for patients with refractory disease prior to colectomy. The follow-up time afforded by infliximab treatment may allow for more accurate classification of the disease in a significant proportion of patients whose colitis has indeterminate features at initial presentation.

A follow-up study of Toxoplasma gondii infection in southern Brazil.

PURPOSE: To understand better the natural history of ocular toxoplasmosis by reexamining a well-characterized population in Southern Brazil. METHODS: Ophthalmological examination and serologic tests for Toxoplasma gondii infection were performed in 1997 on 383 individuals who had undergone the same evaluation in 1990. RESULTS: Of 109 seronegative subjects in 1990, 21 (19.3%) became seropositive by 1997, and 2 (1.5% of previously seronegative patients; 9.5% of those known to have seroconverted) developed ocular toxoplasmosis. Seroconversion occurred more frequently in individuals under 17 years of age (16 of 46 patients, 34.8%) than in those greater than 17 years of age (5 of 63 patients, 7.9%; p = 0.002). Of 131 seropositive individuals who did not have ocular lesions in 1990, 11 (8.3%) had typical toxoplasmic lesions in 1997. Of the 13 individuals with non-specific hyperpigmented small retinal lesions in 1990, 3 (23%) presented with typical lesions in 1997. CONCLUSIONS: Acquired T. gondii infection can result in late development of ocular lesions. Small, non-specific hyperpigmented retinal lesions may represent sites of T. gondii infection in seropositive individuals.

Prospective and controlled study of ophthalmopathy after radioiodine therapy for Graves' hyperthyroidism.

The effects of radioiodine (131I) therapy for hyperthyroidism on the ocular process of Graves' disease is controversial. In order to evaluate the outcome of ophthalmopathy after radioiodine therapy for thyrotoxicosis we studied prospectively 30 Graves' hyperthyroid patients, 22 submitted to radioiodine (131I) treatment (group A) and 8 treated with antithyroid drugs (group B). All patients were evaluated by clinical ophthalmologic examination, and ocular proptosis (OP) was measured with both a Hertel exophthalmometer (HE) and computed tomography (CT) before and 4 to 7 months after therapy. No statistical difference was obtained between pre- and post-treatment OP measurements in each eye in either group, and we did not observe worsening in the ophthalmopathy of patients treated with drugs or radioiodine. After therapy, there was an improvement in the clinical signs of ophthalmopathy in 59% of group A and in 37.5% of group B patients. We found a significant correlation between OP measured by HE and by CT. CT findings showed an increase in orbital fat and/or muscle thickening in all patients at baseline, proving to be a useful procedure for ophthalmologic diagnosis in doubtful cases. No patient in either group developed hypothyroidism or elevated TSH levels during the study period; this may explain our good results in the evolution of Graves' ophthalmopathy after treatment with 131I and antithyroid drugs. Euthyroidism seems to be an important factor in the outcome of ophthalmopathy after therapy, whatever the mode of treatment chosen to achieve it.

Ocular manifestations in the acute phase of leptospirosis.

OBJECTIVES: To study the ocular manifestations during the acute phase of leptospirosis and their incidence in hospitalized patients due to systemic complications, and to verify the importance of routine ophthalmologic care in these cases. METHODS: Twenty-one patients, 20 males and 1 female, with clinical and laboratory (ELISA IgM) diagnoses of leptospirosis were subjected to ophthalmologic examination. RESULTS: We observed conjunctival hyperemia in 18 patients (85.7%), increased retinal venous caliber in 12 patients (57.1%), optic disc redness in 12 patients (57.1%), subconjunctival hemorrhage in four patients (19.0%), optic disc edema in one patient (4.8%), retinal vasculitis in one patient (4.8%), retinal hemorrhage in one patient (4.8%), hard exudates in one patient (4.8%), and papillitis in one patient (4.8%). No anterior chamber reaction was found. CONCLUSIONS: We observed a high incidence of several ocular manifestations in the acute phase of leptospirosis. Despite the systemic severity and high incidence of ocular disorders in the acute phase of leptospirosis, the short-term visual outcome of these patients was good.

Vogt-Koyanagi-Harada's disease in Brazil.

A retrospective analysis of 33 patients with Vogt-Koyanagi-Harada disease (VKH disease) seen in São Paulo, Brazil, from 1976 to 1985 at a uveitis referral clinic revealed that VKH disease represents 2.5% of the total uveitis cases seen. All cases were bilateral, 30% being men and 70% women. The ethnic distribution was the following: 60% white (with variable Indian or black extraction), 24% darkly pigmented, 9% Orientals (Sansei, third-generation Japanese) and 6% black. The frequency among Orientals was 7 times higher than what would be expected according to the relative frequency of Japanese in the Brazilian population. The age distribution at the onset of the disease was as follows: 12% less than 20 years of age, 60% between 20 and 40 years of age and 27% over 40 years of age. The disease was classified into 3 types with variable extraocular signs. Type I disease was present in 24% of the patients, type II in 51% and type III disease in 24% of the patients. Cataract was present in 40% of the cases and glaucoma was present in 9%. No correlation was found between sex, age at onset, race, type of extraocular involvement and number of extraocular manifestations in considering either visual status or visual prognosis. All patients were treated with systemic steroids. Most of them also received cytotoxic immunosuppressive agents. In this uncontrolled clinical study cytotoxic drug-treated patients seemed to have a better clinical course.

Behçet's disease in Brazil--a review of 49 cases with emphasis on ophthalmic manifestations.

At the Uveitis Clinic of the Department of Ophthalmology of the Escola Paulista de Medicina, São Paulo, Brazil, Behçet's disease was diagnosed in 49 patients, representing 2.0% of the total uveitis cases attended during the 16-year period from February 1974 to June 1990. Of these, 71% were men. The age of onset of the disease was between 9 and 61 years, with a mean of 29.6 years. The ethnic distribution was the following: 76% Caucasian, 14% darkly pigmented, 8% Mongoloid and 2% Negroid. HLA-B5 was found in 45.5% of the 11 Caucasian patients typed. In 34.5% of the cases, the ocular attack was the initial manifestation, alone or associated with other symptoms. Oral aphthae were recorded in 98% of the patients, genital ulcers in 55.1%, and skin lesions in 51%. Joint involvement was present in 44.9%, neurologic symptoms were evidenced in 3 patients, and 2 patients had major vascular involvement. The mean interval between the first manifestation of the disease and the onset of ocular involvement was 3.1 years, with a range of 4 months to 14 years. The interval between affections of both eyes ranged from 0 to 2 years; in 38.4% of the cases it occurred within one month. Anterior and posterior segment involvement was seen in 85.7% of the patients. Hypopyon was observed in 34.7% of the cases. Seven patients did not present iridocyclitis at any time in the course of their disease. We did not see any cases with only anterior segment involvement.(ABSTRACT TRUNCATED AT 250 WORDS)

[Ophthalmological manifestations in AIDS: evaluation of 445 patients in one year]. / Achados oftalmológicos em AIDS: avaliação de 445 casos atendidos em um ano.

The authors prospectively evaluated 445 HIV positive patients for the presence of ophthalmological manifestations. PURPOSE--To evaluate patients HIV positive with or without AIDS and correlate the data with the ocular findings mentioned in the literature. METHODS--445 HIV positive patients (66% with AIDS) were evaluated in one year at the Paulista School of Medicine, São Paulo Hospital, Brazil. There was a predominance of males (87%) and homosexuals (58.2%). RESULTS--Of the 445 patients, 52% presented ocular findings secondary to HIV infection at the first examination. The diagnosis included: CMV retinitis (25%), ocular toxoplasmosis (8.5%), herpes retinitis (3.6%), papilledema (2.2%), optic atrophy (1.6%), phthisis bulbi (1.5%), multifocal choroiditis (1.2%), retinal hemorrhages (0.9%), syphilitic uveitis (0.6%) and central vein occlusion (0.2%). CONCLUSION--The incidence of ophthalmic manifestations of AIDS in Brazil is similar to that found in the international literature. We found though a higher incidence of ocular toxoplasmosis than that in other countries. No ocular pneumocystosis was presents in the population evaluated by us.

The incidence and risk factors for developing depression after being diagnosed with inflammatory bowel disease: a cohort study.

BACKGROUND: Studies have found that depression is more frequent in patients with inflammatory bowel disease (IBD) than the general population. Clinicians are now trying to pinpoint risk factors for psychological impairment in the IBD population. AIMS: To examine the demographic and phenotypic variables associated with the development of depression among a diverse cohort of IBD patients. We also sought to describe psychotropic therapy prescribed to IBD patients. METHODS: We conducted a retrospective cohort study including patients with Crohn's disease (CD) and ulcerative colitis (UC) without a prior psychiatric diagnosis and followed in the gastroenterology clinics of the private university hospital and public safety net hospital at a large academic centre in Miami (Florida). Predictive variables included demographic characteristics, IBD phenotype, exposure to IBD medications, history of a surgical stoma or seton placement, extra-intestinal manifestations, laboratory indices, aggressive disease and disease activity (based on imaging and endoscopic parameters). Proportional hazard regression models and stepwise Cox regression analysis were used for statistical analysis. RESULTS: Independent predictors of depression were female gender [HR: 1.3 (95% CI: 1.1-1.7), P = 0.01], aggressive disease [HR: 1.4 (95% CI: 1.02-1.9), P = 0.03] and active disease [HR: 1.5 (95% CI: 1.1-2.0), P = 0.04]. In the group that did develop a depressive disorder, 65% received pharmacologic therapy with one or more psychotropic agents. CONCLUSIONS: We found female gender, aggressive disease and increased endoscopic/radiological activity to be independently associated with the development of depression in inflammatory bowel disease.

Early clinical experience with adalimumab in treatment of inflammatory bowel disease with infliximab-treated and naïve patients.

BACKGROUND: Adalimumab, at an induction dose of 160/80 mg followed by 40 mg every other week is approved for treatment of refractory Crohn's disease (CD) and for patients with loss of response to infliximab. AIM: To evaluate the indications for adalimumab, the proportion of inflammatory bowel disease patients who require dose escalation and to identify whether this strategy is effective in inducing or maintaining remission. METHODS: Patients prescribed adalimumab for CD were identified and included for analysis, if they had follow-up of at least 6 weeks. Adalimumab dose was escalated if patients had return of symptoms prior to next dose. Clinical judgment was used to determine severity of disease. A second GI physician confirmed disease severity as determined by the first physician. RESULTS: A total of 48 out of 60 patients met inclusion criteria. Adalimumab was used to treat CD in 47/48 (98%) and ulcerative colitis in one (2%). Most patients had moderate 30/48 (63%) or severe 17/48 (35%) disease. Prior infliximab exposure was present in 42/48 (88%). Adalimumab dose escalation occurred in 14/48 (29%) within an average time of 2.2 months (s.d. 1.5 months). A majority of patients who required dose escalation, nine of 14 (64%) did not improve clinically. Steroids could be discontinued in three of 16 (18.8%). Clinical improvement was noted in 21/48 (43.8%) and one of 48 (2%) patients achieved clinical remission. Adverse drug reactions necessitated drug discontinuation in four of 48 (8%) of patients. CONCLUSIONS: This retrospective review from a single academic medical centre suggests that a minority of patients, who cannot be maintained on 40 mg every other week, of adalimumab benefit from an increased dose. This suggests the need for a treatment with an alternative mode of action in anti-TNF failures.

NSAID-induced antral ulcers are associated with distinct changes in mucosal gene expression.

BACKGROUND: The basis for individual variation in gastroduodenal vulnerability to NSAIDs is not well understood. AIM: To assess whether a gene expression signature is associated with susceptibility to gastroduodenal ulcerations. METHODS: Twenty-five Helicobacter pylori negative adults were treated for 7 days with naproxen 500 mg b.d. Subjects underwent baseline and post-treatment endoscopy, during which biopsies were taken from antrum and duodenum. RNA extraction and cDNA synthesis were performed, followed by PCR of 23 genes relevant to mucosal injury and repair. Fold changes in gene expression were compared between subjects who developed ulcers and those who did not. RESULTS: Compared with subjects who did not develop ulcers (n = 18), subjects who developed antral ulcers (n = 7) had significantly greater mucosal up-regulation of interleukin-8 [Fold change = 33.5 (S.E.M. = 18.5) vs. -7.7 (3.2)] and of cyclo-oxygenase-2 [2.3 (1.7) vs. -10.8 (2.2)]. Conversely, non-ulcer subjects had significantly greater up-regulation of toll-like receptor-4, cyclo-oxygenase-1 and hepatocyte growth factor [14.0 (2.2) vs. -0.8 (1.0), 9.8 (2.4) vs. 0.0 (0.7) and 8.2 (2.6) vs. -2.2 (0.3) respectively]. CONCLUSIONS: NSAID-induced antral ulcers are associated with a specific pattern of gastroduodenal mucosal gene expression. These patterns may provide an insight into the molecular basis of individual susceptibility to mucosal injury.

Role of Toll-like receptors in gastrointestinal malignancies.

Mounting evidence supports the tenet that innate immune responses to luminal microbes participate in the development of gastrointestinal malignancies. The gastrointestinal tract is relatively unique in that it has evolved in the presence of diverse enteric microflora. Intestinal flora is required to develop a normal adaptive immune response in the periphery. With the characterization of the innate immune system, we have begun to understand the adaptations the intestine has made to the microbiota. The interaction between the microbiota and the intestinal mucosa through Toll-like receptors (TLRs) is required to maintain intestinal homeostasis. In particular, intestinal epithelial cells and lamina propria mononuclear cells such as antigen-presenting cells and T cells must respond to breaches in the mucosal barrier by activating TLR-dependent pathways that result in increased epithelial proliferation, wound healing and recruitment of acute inflammatory cells. In the setting of chronic inflammation such as Helicobacter pylori (H. pylori) infection in the stomach or idiopathic inflammatory bowel disease, the process of repair may eventually result in carcinogenesis. The following review highlights human and animal data that support a role for innate immune responses and TLRs specifically in promoting gastrointestinal malignancies. Candidate pathways linking TLRs to gastrointestinal malignancies include activation of nuclear factor-kappaB and cyclooxygenase-2. Studying the link between innate immune signaling and gastrointestinal malignancies offers the possibility to identify novel ways to both prevent and treat gastrointestinal cancer.

TLR4 signalling in the intestine in health and disease.

The colonic epithelium is lined along its apical membrane with approximately 10(14) bacteria/g of tissue. Commensal bacteria outnumber mammalian cells in the gut severalfold. The reason for this degree of commensalism probably resides in the recent recognition of the microbiome as an important source of metabolic energy in the setting of poorly digestible nutrients. As in many themes in biology, the host may have sacrificed short-term benefit, i.e. nutritional advantages, for long-term consequences, such as chronic inflammation or colon cancer. In the present review, we examine the role of TLR (Toll-like receptor) signalling in the healthy host and the diseased host. We pay particular attention to the role of TLR signalling in idiopathic IBD (inflammatory bowel disease) and colitis-associated carcinogenesis. In general, TLR signalling in health contributes to homoeostatic functions. These include induction of antimicrobial peptides, proliferation and wound healing in the intestine. The pathogenesis of IBD, ulcerative colitis and Crohn's disease may be due to increased TLR or decreased TLR signalling respectively. Finally, we discuss the possible role of TLR signalling in colitis-associated neoplasia.

The effect of adrenalectomy on Fos expression in vasopressinergic and oxytocinergic neurons in response to stress in the rat.

This study evaluated the responses of vasopressin (AVP) and oxytocin (OT) neurons to alterations in hypothalamo-pituitary axis activity by adrenalectomy (ADX) or after restraint stress compared with basal conditions. Wistar male rats were perfuse-fixed by cardiac perfusion under anesthesia 3 h, 1, 3 and 14 days after ADX or Sham surgery. Coronal hypothalamic sections were used for evaluation of Fos, AVP and OT expression by immunohistochemistry. Under basal conditions and after stress, Fos-AVP double labeling showed no difference in the magnocellular subdivisions of the paraventricular nuclei (PVN) or in the supraoptic nuclei (SON), suggesting that the magnocellular AVP system is unlikely to contribute to ACTH secretion after restraint in both Sham and ADX rats. Fos-AVP double labeling in the parvocellular medial paraventricular nucleus (PaMP) in ADX groups was increased after 3 h in basal conditions, and in all periods after restraint stress. There were no differences between Sham and ADX groups in Fos-OT double labeling in any subdivision of the PVN; however, in the SON, the number of Fos-OT double labeled cells was increased at all time-points after stress in the ADX group. Fos expression was increased in the PaMP after 3 h and after restraint stress in the Sham and ADX groups, especially in the ADX group. In conclusion, Fos expression in different cell populations of the PVN can be differentially regulated by short- and long-term absence of glucocorticoid negative feedback and also by stress-related excitatory and/or inhibitory neural inputs. The Fos-AVP double labeling findings in the PaMP also indicate a minor participation of these vasopressinergic neurons in the regulation of the HPA axis after ADX.