Stefan J. Urbanski

Altered Balance Between Matrix Metalloproteinases and Their Inhibitors in Experimental Biliary Fibrosis

A rat model of common bile duct ligation (BDL)-induced hepatic fibrosis was used to assess the expression and activities of collagen-degrading proteinases and their inhibitors during the progression of fibrosis. Expression of four members of the matrix metalloproteinase (MMP) family (MMP-2/gelatinase A, MMP-3, MMP-9/gelatinase B, and MMP-13) and three tissue inhibitors of metalloproteinases-1, -2, and -3 (TIMP-1, TIMP-2, and TIMP-3) were evaluated by Northern blot analysis of RNA from liver tissue isolated at 0, 2, 5, 10, 20, and 30 days after either a BDL or sham operation. In addition, we analyzed free gelatinase and TIMP activities by zymography and reverse zymography, respectively. We found that the proteolytic activities of MMP-2 and MMP-9 increased by 2 days after ligation, reached maximal levels at day 10, and remained high through the study period, whereas the gelatinolytic activities in plasma were unchanged. The increase in gelatinase activities was accompanied by an increase in the TIMP mRNA transcripts. TIMP-1 transcripts appeared at day 2, increased until day 10, and remained elevated throughout the study period. TIMP-2 and TIMP-3 transcripts become detectable on day 10 and remained stable afterwards. No corresponding increase in TIMP protein activity was detected by reverse zymography. This appears to result from the formation of TIMP/MMP complexes. These findings indicate a likely surplus in the BDL model of fibrosis of free gelatinases as compared with the TIMPs. Thus, excessive TIMP production is not a sufficient explanation for the observed extracellular matrix accumulation, but complex changes in the local MMP/TIMP balance may underlie the pathomechanisms of fibrosis.

The burden of large and small duct primary sclerosing cholangitis in adults and children: a population-based analysis.

OBJECTIVES:The epidemiology of primary sclerosing cholangitis (PSC) has been incompletely assessed by population-based studies. We therefore conducted a population-based study to determine: (a) incidence rates of large and small duct PSC in adults and children, (b) the risk of inflammatory bowel disease on developing PSC, and (c) patterns of clinical presentation with the advent of magnetic resonance cholangiopancreatography (MRCP).METHODS:All residents of the Calgary Health Region diagnosed with PSC between 2000 and 2005 were identified by medical records, endoscopic, diagnostic imaging, and pathology databases. Demographic and clinical information were obtained. Incidence rates were determined and risks associated with PSC were reported as rate ratios (RR) with 95% confidence intervals (CI).RESULTS:Forty-nine PSC patients were identified for an age- and gender-adjusted annual incidence rate of 0.92 cases per 100,000 person-years. The incidence of small duct PSC was 0.15/100,000. In children the incidence rate was 0.23/100,000 compared with 1.11/100,000 in adults. PSC risk was similar in Crohns disease (CD; RR 220.0, 95% CI 132.4–343.7) and ulcerative colitis (UC; RR 212.4, 95% CI 116.1–356.5). Autoimmune hepatitis overlap was noted in 10% of cases. MRCP diagnosed large duct PSC in one-third of cases. Delay in diagnosis was common (median 8.4 months). A minority had complications at diagnosis: cholangitis (6.1%), pancreatitis (4.1%), and cirrhosis (4.1%).CONCLUSIONS:Pediatric cases and small duct PSC are less common than adult large duct PSC. Surprisingly, the risk of developing PSC in UC and CD was similar. Autoimmune hepatitis overlap was noted in a significant minority of cases.

Increased gastrointestinal permeability is an early lesion in the spontaneously diabetic BB rat

The BB rat spontaneously develops autoimmune diabetes. Feeding these animals a hydrolyzed casein diet significantly reduces the incidence of this disease, suggesting that a dietary antigen is involved in the pathogenesis of this disease. In other syndromes associated with luminal antigens, including celiac and Crohns disease, increased intestinal permeability has been suggested to play an etiological role. Therefore, the objective of this study was to evaluate whether increased permeability was also present in BB rats before disease development. By measuring gastrointestinal permeability, in animals on a regular or hydrolyzed casein diet, we were able to demonstrate that increased gastric and small intestinal permeability appeared before the development of both insulitis and clinical diabetes. Although hydrolysis of dietary protein significantly reduced the incidence of diabetes, it did not alter the small intestinal permeability abnormality, suggesting that this is an early event. Increased permeability appears to have an early role in the genesis of several immunological diseases and may represent a common event in these diseases.The BB rat spontaneously develops autoimmune diabetes. Feeding these animals a hydrolyzed casein diet significantly reduces the incidence of this disease, suggesting that a dietary antigen is involved in the pathogenesis of this disease. In other syndromes associated with luminal antigens, including celiac and Crohns disease, increased intestinal permeability has been suggested to play an etiological role. Therefore, the objective of this study was to evaluate whether increased permeability was also present in BB rats before disease development. By measuring gastrointestinal permeability, in animals on a regular or hydrolyzed casein diet, we were able to demonstrate that increased gastric and small intestinal permeability appeared before the development of both insulitis and clinical diabetes. Although hydrolysis of dietary protein significantly reduced the incidence of diabetes, it did not alter the small intestinal permeability abnormality, suggesting that this is an early event. Increased permeability appears to have an early role in the genesis of several immunological diseases and may represent a common event in these diseases.

Microscopic Colitis–Defining Incidence Rates and Risk Factors: A Population-Based Study

BACKGROUND & AIMS The burden and determinants of microscopic colitis (MC) in North America are inadequately defined. We determined the incidence rate of and risk factors for MC in a well-defined North American population. METHODS A population-based cohort study was conducted between April 1, 2002, and March 31, 2004. All adults with a pathologic diagnosis of MC were identified and comprehensive chart review was undertaken to confirm the diagnosis and identify risk factors. Category-specific risks for developing MC were reported as rate ratios (RRs) with exact 95% confidence intervals (CIs). RESULTS MC was identified in 164 individuals for an annual incidence rate of 10.0 per 100,000 person-years (lymphocytic colitis, 5.4; collagenous colitis, 4.6 per 100,000). Patients older than the age of 65 were more than 5 times more likely to develop MC (RR, 5.6; 95% CI, 4.0-7.7). Women were at higher risk of acquiring MC for both collagenous colitis (RR, 3.44; 95% CI, 2.07-5.97) and lymphocytic colitis (RR 6.29; 95% CI, 3.21-13.74). Elderly women with a history of malignancy were associated with a higher risk of MC (RR, 3.59; 95% CI, 1.68-7.01), as were patients with celiac disease (RR, 7.9; 95% CI, 4.0-14.2) and hypothyroidism (RR, 6.1; 95% CI, 3.5-10.0). CONCLUSIONS This was a large population-based cohort study of MC and our incidence rates were consistent with previously reported population-based studies in North America and Europe. An increased incidence of MC was observed in several disease states with the novel finding of an increased risk of MC with malignancy.

Mycophenolate mofetil for the treatment of autoimmune hepatitis in patients refractory to standard therapy.

There are limited therapeutic options available for patients with autoimmune hepatitis in whom conventional treatment fails. A case series of five patients unresponsive to or unable to take azathioprine, 6-mercaptopurine or corticosteroids who were treated with mycophenolate mofetil (MMF) is reported. While on MMF, alanine aminotransferase normalized or remained normal in all patients. MMF had a steroid-sparing effect and histological remission was demonstrated in one patient after seven months of MMF. One patient experienced an uncomplicated episode of pyelonephritis. In conclusion, MMF can effectively induce and maintain remission in refractory autoimmune hepatitis patients.

Successful treatment of refractory type 1 autoimmune hepatitis with methotrexate

BACKGROUND/AIMS Autoimmune hepatitis is a heterogeneous disorder that typically responds to glucocorticoids with or without azathioprine. Treatment options for patients not responding to standard therapy are limited. METHODS We describe a 52-year-old female who presented with jaundice, marked elevation in liver enzymes, positive antinuclear antibody and a liver biopsy consistent with autoimmune hepatitis. Liver enzymes did not normalize with prednisone alone. When azathioprine was added, the disease flared. The patient refused cyclosporine. Methotrexate 7.5 mg po per week resulted in normalization of liver enzymes, improved liver histology, and has maintained remission with a steroid-sparing effect. RESULTS/CONCLUSION In this patient methotrexate was used successfully to treat type 1 autoimmune hepatitis. This suggests that methotrexate may have a role in treatment of autoimmune hepatitis refractory to standard therapy.

Lack of Chemokine Receptor CCR5 Promotes Murine Fulminant Liver Failure by Preventing the Apoptosis of Activated CD1d-Restricted NKT Cells

Fulminant liver failure (FLF) consists of a cascade of events beginning with a presumed uncontrolled systemic activation of the immune system. The etiology of FLF remains undefined. In this study, we demonstrate that CCR5 deficiency promotes the development of acute FLF in mice following Con A administration by preventing activated hepatic CD1d-restricted NKT cells (but not conventional T cells) from dying from activation-induced apoptosis. The resistance of CCR5-deficient NKT cells from activation-induced apoptosis following Con A administration is not due to a defective Fas-driven death pathway. Moreover, FLF in CCR5-deficient mice also correlated with hepatic CCR5-deficient NKT cells, producing more IL-4, but not IFN-γ, relative to wild-type NKT cells. Furthermore, FLF in these mice was abolished by IL-4 mAb or NK1.1 mAb treatment. We propose that CCR5 deficiency may predispose individuals to the development of FLF by preventing hepatic NKT cell apoptosis and by regulating NKT cell function, establishing a novel role for CCR5 in the development of this catastrophic liver disease that is independent of leukocyte recruitment.

Prevalence, risk factors and causes of discordance in fibrosis staging by transient elastography and liver biopsy

Background and aims: Liver stiffness measurement (LSM) by transient elastography (TE) is widely used for the noninvasive assessment of fibrosis. Our objectives were to examine the prevalence, risk factors and causes of discordance between fibrosis estimated by TE and liver biopsy.

Technical considerations for prophylactic mastectomy in patients at high risk for breast cancer

A study of 5 patients and 10 mastectomy specimens was performed to identify the extent of surgery necessary to completely remove all breast tissue in patients having prophylactic mastectomies. A standard total mastectomy performed for breast cancer was shown to frequently leave breast tissue within the superficial pectoralis major muscle and the lower skin flap. Frozen section analysis of margins was found to be essential to clear the axillary extension of the breast and lower skin flap in particular. The value of more extensive surgery to remove all glandular elements of the breast in the high-risk patient remains to be demonstrated.

Rituximab for the treatment of patients with autoimmune hepatitis who are refractory or intolerant to standard therapy

BACKGROUND Although most patients with autoimmune hepatitis (AIH) respond to treatment with prednisone and⁄or azathioprine, some patients are intolerant or refractory to standard therapy. Rituximab is an anti-CD20 monoclonal antibody that depletes B cells and has demonstrated efficacy in other autoimmune conditions. AIMS To evaluate the safety and efficacy of rituximab in patients with refractory AIH in an open-label, single-centre pilot study. METHODS Six patients with definite, biopsy-proven AIH who failed prednisone and azathioprine treatment received two infusions of rituximab 1000 mg two weeks apart and were followed for 72 weeks. RESULTS Rituximab was well tolerated with no serious adverse events. By week 24, mean (± SD) aspartate aminotransferase (AST) levels had significantly improved (90.0±23.3 U⁄L versus 31.3±4.2 U⁄L; P=0.03) and mean immunoglobulin G levels had fallen (16.4±2.0 g⁄L versus 11.5±1.1 g⁄L; P=0.056). The prednisone dose was weaned in three of four subjects, with one subject flaring after steroid withdrawal. Inflammation grade improved in all four subjects who underwent repeat liver biopsy at week 48. Regulatory T cell levels examined by FoxP3 immunohistochemistry paralleled inflammatory activity and did not increase on follow-up biopsies. There was no significant change in serum chemokine or cytokine levels from baseline to week 24 (n=5), although interferon-gamma-induced protein 10 levels improved in three of five subjects. CONCLUSIONS Rituximab was safe, well tolerated and resulted in biochemical improvement in subjects with refractory AIH. These results support further investigation of rituximab as a treatment for AIH.