In vitro-induced antibody production in chronic hepatitis C virus infection

The objectives of the present study were to assess the in vitro-induced anti-hepatitis C virus (HCV) antibody production (IVIAP) in relation to the clinical, biochemical, virologic and histologic variables of patients with HCV infection. The study included 57 patients (60% males) with HCV infection (anti-HCV and HCV-RNA positive). Alanine aminotransferase (ALT) was elevated in 89% of the patients. Mean viral load was 542,241 copies/ml and histology of the liver showed chronic hepatitis in 27/52 (52%) and cirrhosis in 11/52 (21%) patients. IVIAP levels were determined by immunoenzymatic assay at median absorbance of 0.781 at 450 nm. IVIAP was negative in 14% of the patients. When groups with IVIAP levels above and below the median were compared, high IVIAP levels were associated with the male sex, elevated ALT levels and more advanced disease stage. After logistic regression analysis, advanced histologic damage to the liver remained as the only independent variable associated with elevated IVIAP levels. Using a receiver operator characteristic curve, the best cut-off level for IVIAP was established (= 1.540), with 71% sensitivity and 94% specificity for the detection of more advanced disease stages (grades 3 and 4). These findings are consistent with the participation of immunological mechanisms in the genesis of the hepatic lesions induced by HCV and indicate that the IVIAP test may be useful as a noninvasive marker of liver damage either alone or in combination with other markers.

Functional dyspepsia: relationship between clinical subgroups and Helicobacter pylori status in Western Turkey

The etiology of functional dyspepsia is not known. The objective of the present study was to determine the characteristics of functional dyspepsia in Western Turkey. We divided 900 patients with functional dyspepsia into three subgroups according to symptoms: ulcer-like (UL), 321 (35.6%), motility disorder-like (ML), 281 (31.2%), and the combination (C) of these symptoms, 298 (33.1%). All patients were submitted to endoscopic evaluation, with two biopsies taken from the cardia and corpus, and four from the antrum of the stomach. All biopsy samples were studied for Helicobacter pylori (Hp) density, chronic inflammation, activity, intestinal metaplasia, atrophy, and the presence of lymphoid aggregates by histological examination. One antral biopsy was used for the rapid urease test. Tissue cagA status was determined by PCR from an antral biopsy specimen by a random sampling method. We also determined the serum levels of tumor necrosis factor-alpha (TNF-alpha) and gastrin by the same method. Data were analyzed statistically by the Kolmogorov-Smirnov test and by analysis of variance. Hp and cagA positivity was significantly higher in the UL subgroup than in the others. The patients in the ML subgroup had the lowest Hp and cagA positivity and Hp density. The ML subgroup also showed the lowest level of Hp-induced inflammation among all subgroups. The serum levels of TNF-alpha and gastrin did not reveal any difference between groups. Our findings show a poor association of Hp with the ML subgroup of functional dyspepsia, but a stronger association with the UL and C subgroups.

Prevalence of celiac disease in Brazilian children of short stature

The aim of the present study was to determine the prevalence of celiac disease in children of short stature and to assess whether some of the routine laboratory examinations performed to determine the cause of short stature could suggest the presence of celiac disease. A total of 106 children of short stature and no gastrointestinal symptoms were studied. An extensive endocrine work-up had been negative for all of them and an additional investigation was performed by measuring the concentration of antiendomysial antibody. Patients who were positive for antiendomysial antibody ( > or = 1:10) or who exhibited IgA deficiency (less than 5 mg/dl) were referred for an endoscopic intestinal biopsy. We detected a pathological titer of antiendomysial IgA in six of these patients. Five of them showed histological abnormalities compatible with celiac disease and one had normal histology and was considered to have potential celiac disease. The prevalence of celiac disease in the population studied was 4.7% (with another 0.9% of the subjects being considered to have potential celiac disease). The children with celiac disease did not differ in any of the parameters tested when compared to those without celiac disease, though they showed an improvement in growth velocity after treatment with a gluten-free diet. We conclude that it is important to test all children with short stature for celiac disease by measuring antiendomysial IgA.

Estrogen and progesterone receptors in human papilloma virus-related cervical neoplasia

Estrogen (ER) and progesterone (PR) receptors in the normal uterine cervix, cervical intraepithelial neoplasia and invasive carcinoma were studied in consecutive samples from Hospital do Câncer, São Paulo, between 1996 and 1997. Tissue was collected by removing a fragment of the tumoral area using a 5-mm diameter biopsy punch, followed by removal of a macroscopically normal area as close as possible from the tumor. Histopathological confirmation was obtained for all specimens analyzed. A total of 24 normal tissues, 17 cases of cervical intraepithelial neoplasia and 7 of invasive carcinomas were studied. The ER/PR ratio was determined by immunohistochemistry using monoclonal antibodies specific for each receptor. Adjacent tissue slides were submitted to generic PCR for human papillomavirus (HPV) DNA detection followed by typing by dot blot hybridization. About half (45.8%) of the tumors were HPV DNA positive while 29.1% of the patients were also HPV positive in their respective normal tissue. ER was negative in the tumoral epithelium of 11 HPV-positive patients (P = 0.04). There was a trend in the ER distribution in normal tissue that was opposite to that from lesions, but it was not statistically significant (P = 0.069). No difference in ER distribution in stromal tissues was observed between HPV-positive and HPV-negative tissues. PR staining was negative in the epithelium of all cases studied. The results obtained from this small number of cases cannot be considered to be conclusive but do suggest that factors related to viral infection affect the expression of these ER/PR cervix receptors.

Dermal dendritic cell number correlates with serum autoantibody titers in Brazilian pemphigus foliaceus patients

Pemphigus foliaceus (PF) is an autoimmune bullous disease endemic in Brazil. Since serum IL-12 is increased in patients with PF and Langerhans cells (LC) produce IL-12, we titrated serum autoantibodies by indirect immunofluorescence, and quantified epidermal dendritic cells, known as LC, and dermal dendritic cells (DC). Biopsies of blistering lesions were obtained from 22 patients, 13 of whom were submitted to biopsy of both injured and of apparently healthy skin. The control groups consisted of skin from 8 cadavers and from 12 women submitted to breast plastic surgery. LC and DC were identified with anti-CD1a antibody and quantified by morphometric analysis. LC number in the lesion and in apparently healthy skin from PF patients was similar to that of both control groups. DC number in the injured skin (median = 0.94 DC/mm basement membrane) was higher than that of the cadaver group (median = 0.13 DC/mm basement membrane). In the 13 patients with biopsies of both injured and apparently healthy skin, LC and DC were present in larger numbers in the lesion. There was a direct correlation between DC number in the lesion of the PF group and serum autoantibody titers. This correlation was not observed for LC number. The increased number of DC in the lesion, as well as its direct correlation with serum autoantibody titers suggest the participation of DC in the pathogenesis of PF. The relationship between increased DC number and IL-12 in PF needs to be clarified.

BCG lymphadenopathy detected in a BCG-vaccinated infant

Large-scale vaccination with BCG, the live attenuated strain of Mycobacterium bovis, is being adopted around the world, although sporadic complications have occurred after the procedure. Lymphadenopathy is not uncommon especially in babies under one year (0.73% of vaccinated infants), but the swelling subsides within 2 months in most cases, with no medical or surgical treatment. Brazil adopted BCG vaccination program earlier in the seventies and by 1995 more than 96% of the infant population received this immunization. We report here the occurrence of lymphadenopathy in a two-year-old child vaccinated with the Brazilian BCG strain. The diagnosis was made using a lymph node biopsy and intestinal aspirates that yielded a positive mycobacterial culture. The isolate was resistant to isoniazid, rifampicin, pyrazinamide and thiophen-2-carbonic acid hydrazide, sensitive to streptomycin, ethambutol, and p-nitrobenzoic acid, and reacted positively to cyclo-serine and negatively to niacin. The pncA gene involved in bacterial activation of pyrazinamide contains in M. bovis a point mutation that renders pyrazinamidase unable to catalyze drug activation. Therefore, this polymorphism is a good option for developing methods to differentiate M. bovis and M. tuberculosis. Taking advantage of this difference we further analyzed the isolates by single-stranded conformation polymorphism electrophoresis of DNA following PCR of the pncA gene. The isolate identity was confirmed by RFLP electrophoretic analysis of the amplified fragment following Eco065I digestion, which selectively cleaves M. tuberculosis DNA. From this result it is proposed that RFLP of pncA gene represents an alternative for differential diagnosis of M. bovis.

Preservation of graft function in low-risk living kidney transplant recipients treated with a combination of sirolimus and cyclosporine

The use of sirolimus (SRL) in combination with full doses of cyclosporin A (CsA) results in reduced one-year kidney allograft function, which is associated with shorter long-term allograft survival. We determined the effect of reduced CsA exposure on graft function in patients receiving SRL and prednisone. Ninety recipients of living kidney transplants receiving SRL (2 mg/day, po) were compared to 35 recipients receiving azathioprine (AZA, 2 mg kg-1 day-1, po). All patients also received CsA (8-10 mg kg-1 day-1, po) and prednisone (0.5 mg kg-1 day-1). Efficacy end-point was a composite of biopsy-confirmed acute rejection, graft loss, or death at one year. Graft function was measured by creatinine, creatinine clearance, and graft function deterioration between 3 and 12 months (delta1/Cr). CsA concentrations in patients receiving SRL were 26% lower. No differences in one-year composite efficacy end-point were observed comparing SRL and AZA groups (18 vs 20%) or in the incidence of biopsy-proven acute rejection (14.4 and 14.3%). There were no differences in mean ± SD creatinine (1.65 ± 0.46 vs 1.60 ± 0.43 mg/dl, P = 0.48) or calculated creatinine clearances (61 ± 15 vs 62 ± 13 ml/min, P = 0.58) at one year. Mean ± SD delta1/Cr (-11 ± 17 vs -14 ± 15%, P = 0.7) or the percentage of patients with >20% (26 vs 31%, P = 0.6) or >30% delta1/Cr (19 vs 17%, P = 1) did not differ between the two groups. The use of 2-mg fixed oral doses of SRL and reduced CsA exposure was effective in preventing acute rejection and preserving allograft function.

Comparative study of IgA nephropathy with and without crescents

Glomerular crescents were analyzed as a prognostic factor in retrospectively reviewed data from 144 patients with biopsy-proven IgA nephropathy. Crescents were found in 26 (18%) patients, and detected in 2 to 100% of glomeruli in each specimen. In 5% of the patients more than 50% of the glomeruli were affected. Thirty patients with IgA nephropathy without crescents were studied as a control group. Mean age was 30.3 ± 9.4 and 30.2 ± 12.0 years for the patients with and without crescents, respectively, and males prevailed in both groups. The length of follow-up was 23.2 ± 41.6 months for patients with crescents and 29.3 ± 35.3 months for patients without crescents. Eighty percent of the patients with crescents were hypertensive, compared to 27% of the non-crescent control group (P < 0.05). Mean serum creatinine at the time of diagnosis was 3.9 ± 2.9 and 1.9 ± 2.1 mg/dl for the patients with and without crescents, respectively. Initial urinary protein excretion was higher in patients with crescents (4.6 ± 3.5 vs 1.2 ± 0.9 g/day; P < 0.05). At the end of follow-up 17 patients (77.3%) from the crescent group and 3 (11.1%) from the non-crescent group had end-stage renal disease (P < 0.0001). The presence of crescents was associated with higher levels of initial serum creatinine and urinary protein excretion, and a higher frequency of hypertension and progression to end-stage renal disease.

One-year follow-up of the effects of sildenafil on pulmonary arterial hypertension and veno-occlusive disease

We hypothesized that chronic oral administration of the phosphodiesterase-5 inhibitor sildenafil could improve the exercise capacity and pulmonary hemodynamics in patients with pulmonary arterial hypertension (PAH) on the basis of previous short-term studies. We tested this hypothesis in 14 subjects with PAH, including seven patients with the idiopathic form and seven patients with atrial septal defects, but no other congenital heart abnormalities. Patients were subjected to a 6-min walk test and dyspnea was graded according to the Borg scale. Pulmonary flow and pressures were measured by Doppler echocardiography. Patients were given sildenafil, 75 mg orally three times a day, and followed up for 1 year. Sildenafil therapy resulted in the following changes: increase in the 6-min walk distance from a median value of 387 m (range 0 to 484 m) to 462 m (range 408 to 588 m; P < 0.01), improvement of the Borg dyspnea score from 4.0 (median value) to 3.0 (P < 0.01), and increased pulmonary flow (velocity-time integral) from a median value of 0.12 (range 0.08 to 0.25) to 0.23 (range 0.11 to 0.40; P < 0.01) with no changes in pulmonary pressures. In one patient with pulmonary veno-occlusive disease diagnosed by a lung biopsy, sildenafil had a better effect on the pulmonary wedge pressure than inhaled nitric oxide (15 and 29 mmHg, respectively, acute test). He walked 112 m at baseline and 408 m at one year. One patient died at 11 months of treatment. No other relevant events occurred. Thus, chronic administration of sildenafil improves the physical capacity of PAH patients and may be beneficial in selected cases of veno-occlusive disease.

Serum laminin, type IV collagen and hyaluronan as fibrosis markers in non-alcoholic fatty liver disease

Hepatic fibrosis in patients with non-alcoholic fatty liver disease is associated with progression of the disease. In the present study, we analyzed the discriminative ability of serum laminin, type IV collagen and hyaluronan levels to predict the presence of fibrosis in these patients. In this preliminary report, we studied 30 overweight patients divided into two groups according to the absence (group I, N = 19) or presence (group II, N = 11) of fibrosis in a liver biopsy. Triglycerides, aspartate aminotransferase, alanine aminotransferase, gamma-glutamyltranspeptidade, hyaluronan (noncompetitive fluoroassay), type IV collagen, and laminin (ELISA) were determined. Group II presented significantly higher mean laminin, hyaluronan, type IV collagen, and aspartate aminotransferase values, which were due to the correlation between these parameters and the stage of fibrosis in the biopsy (Spearman's correlation coefficient, rS = 0.65, 0.62, 0.53, and 0.49, respectively). Analysis of the ROC curve showed that laminin values >282 ng/ml were those with the best diagnostic performance, with 87% accuracy. Association of laminin with type IV collagen showed improvement in the positive predictive value (100%), but with reduction in diagnostic sensitivity (64%). When compared with the criteria of Ratziu et al. [Gastroenterology (2000) 118: 1117-1123] for the diagnosis of septal fibrosis, laminin values presented a better diagnostic accuracy (83 vs 70%). Determination of extracellular matrix components in serum, especially of laminin, may identify patients with non-alcoholic fatty liver disease and fibrosis and these components may be used as indicators for liver biopsy in these patients.

Hepatitis C virus infection, cryoglobulinemia, and peripheral neuropathy: a case report

Hepatitis C virus (HCV) is essentially hepatotropic but its manifestations can extend beyond the liver. It can be associated with autoimmune diseases, such as mixed cryoglobulinemia, membranoproliferative glomerulonephritis, autoimmune thyroiditis, and lymphoproliferative disorders. The mechanisms that trigger these manifestations are not completely understood. We describe a 48-year-old man with chronic HCV infection (circulating HCV RNA and moderate hepatitis as indicated by liver biopsy), cryoglobulinemia, and sensory and motor peripheral neuropathy. The diagnosis of multineuropathy was confirmed by clinical examination and electromyographic tests. A nerve biopsy revealed an inflammatory infiltrate in the perineurial space and signs of demyelination and axonal degeneration. The patient had no improvement of neurological symptoms with the use of analgesics and neuro-modulators. He was then treated with interferon-alpha (3 million units subcutaneously, 3 times per week) and ribavirin (500 mg orally, twice a day) for 48 weeks. Six months after the end of therapy, the patient had sustained viral response (negative HCV RNA) and remission of neurological symptoms, but cryoglobulins remained positive. A review of the literature on the pathogenesis and treatment of neurological manifestations associated with HCV infection is presented. This report underscores the need for a thorough evaluation of HCV-infected patients because of the possibility of extrahepatic manifestations. Antiviral treatment with interferon and ribavirin can be effective and should be considered in patients with neurological complications associated with HCV infection.

Improvement of adynamic bone disease after renal transplantation

Low bone remodeling and relatively low serum parathyroid hormone (PTH) levels characterize adynamic bone disease (ABD). The impact of renal transplantation (RT) on the course of ABD is unknown. We studied prospectively 13 patients with biopsy-proven ABD after RT. Bone histomorphometry and bone mineral density (BMD) measurements were performed in the 1st and 12th months after RT. Serum PTH, 25-hydroxyvitamin D, 1,25-dihydroxyvitamin D, and osteocalcin were measured regularly throughout the study. Serum PTH levels were slightly elevated at transplantation, normalized at the end of the third month and remained stable thereafter. Bone biopsies performed in the first month after RT revealed low bone turnover in all patients, with positive bone aluminum staining in 5. In the 12th month, second biopsies were performed on 12 patients. Bone histomorphometric dynamic parameters improved in 9 and were completely normalized in 6, whereas no bone mineralization was detected in 3 of these 12 patients. At 12 months post-RT, no bone aluminum was detected in any patient. We also found a decrease in lumbar BMD and an increase in femoral BMD. Patients suffering from ABD, even those with a reduction in PTH levels, may present partial or complete recovery of bone turnover after successful renal transplantation. However, it is not possible to positively identify the mechanisms responsible for the improvement. Identifying these mechanisms should lead to a better understanding of the physiopathology of ABD and to the development of more effective treatments.

Impact of initial exposure to calcineurin inhibitors on kidney graft function of patients at high risk to develop delayed graft function

We conducted a retrospective analysis of the influence of full doses of calcineurin inhibitors [8-10 mg kg-1 day-1 cyclosporine (N = 80), or 0.2-0.3 mg kg-1 day-1 tacrolimus (N = 68)] administered from day 1 after transplantation on the transplant outcomes of a high-risk population. Induction therapy was used in 13% of the patients. Patients also received azathioprine (2 mg kg-1 day-1, N = 58) or mycophenolate mofetil (2 g/day, N = 90), and prednisone (0.5 mg kg-1 day-1, N = 148). Mean time on dialysis was 79 ± 41 months, 12% of the cases were re-transplants, and 21% had panel reactive antibodies >10%. In 43% of donors the cause of death was cerebrovascular disease and 27% showed creatinine above 1.5 mg/dL. The incidence of slow graft function (SGF) and delayed graft function (DGF) was 15 and 60%, respectively. Mean time to last dialysis and to nadir creatinine were 18 ± 15 and 34 ± 20 days, respectively. Mean creatinine at 1 year after transplantation was 1.48 ± 0.50 mg/dL (DGF 1.68 ± 0.65 vs SGF 1.67 ± 0.66 vs immediate graft function (IGF) 1.41 ± 0.40 mg/dL, P = 0.089). The incidence of biopsy-confirmed acute rejection was 22% (DGF 31%, SGF 10%, IGF 8%). One-year patient and graft survival was 92.6 and 78.4%, respectively. The incidence of cytomegalovirus disease, post-transplant diabetes mellitus and malignancies was 28, 8.1, and 0%, respectively. Compared to previous studies, the use of initial full doses of calcineurin inhibitors without antibody induction in patients with SGF or DGF had no negative impact on patient and graft survival.

PCR identification of Mycobacterium tuberculosis complex in a clinical sample from a patient with symptoms of tuberculous spondylodiscitis

A 42-year-old male complaining of thoracic spine pain was admitted to the hospital for evaluation. An X-ray and computer tomography of the thoracic spine showed spondylodiscitis of the L3 lumbar and L2-L3 intervertebral disk. The tuberculin skin test (PPD) was strongly positive. A radioscopy-guided fine needle aspirate of the affected area was cultured but did not reveal the cause of the disease. Two biopsy attempts failed to reveal the cause of the disease by culturing or by acid-fast-resistant staining (Ziehl Neelsen) of the specimens. A third biopsy also failed to detect the infectious agent by using microbiological procedures, but revealed the presence of a 245-bp amplicon characteristic of the Mycobacterium tuberculosis complex after PCR of the sample. The result demonstrates the efficacy of PCR for the identification of M. tuberculosis in situations in which conventional diagnosis by culturing techniques or direct microscopy is unable to detect the microorganism. Following this result the patient was treated with the antituberculous cocktail composed by rifampicin, pirazinamide and isoniazid during a six-month period. At the end of the treatment the dorsalgia symptoms had disappeared.

Immunoexpression of cyclooxygenase-1 and -2 in ulcerative colitis

Ulcerative colitis (UC) is a disease of the colon and rectum characterized by a nonspecific chronic inflammation mediated by the concerted response of cellular and humoral events. Prostaglandins are synthesized by cyclooxygenase (COX)-1 and -2 and exhibit both pro- and anti-inflammatory activity. To evaluate COX-1 and COX-2 immunoexpression in 42 cases of UC and to correlate it with clinicopathological parameters, COX-1 and COX-2 expression was investigated by the immunohistochemistry method. Only patients with all pertinent clinical and evolutive data as well as with adequate biopsy material were included in the study. Fifteen samples of colorectal adenocarcinoma and 14 of large bowel with no histological changes were used for positive and negative controls, respectively. UC patients showed COX-1 immunoreactivity in epithelial cells in 29% of the cases and in inflammatory cells in 43%. COX-2 positivity in epithelial and inflammatory cells was found in 69% of the samples. The comparison between UC and the control groups revealed that the UC group had significantly more positive cases for COX-1 and COX-2 in inflammatory cells. Immunohistochemistry allowed the identification of COX-1 and COX-2 expression in epithelial and inflammatory cells in UC biopsies. No significant difference between COX-1 and COX-2 immunoreactivity in epithelial and inflammatory cells was observed regarding the clinicopathological parameters. COX-2 presented low expression in normal colon and high expression in colorectal adenocarcinoma. COX-2 might play a role in the pathophysiologic processes of inflammatory bowel disease and the development of neoplasia. Treatment with selective COX-2 inhibitors might be an additional option for therapy.