Advantage of ostarine over raloxifene and their combined treatments for muscle of estrogen-deficient rats.

PURPOSE: Selective androgen (ostarine, OST) and estrogen (raloxifene, RAL) receptor modulators with improved tissue selectivity have been developed as alternatives to hormone replacement therapy. We investigated the combined effects of OST and RAL on muscle tissue in an estrogen-deficient rat model of postmenopausal conditions. METHODS: Three-month-old Sprague Dawley rats were divided into groups: (1) untreated non-ovariectomized rats (Non-OVX), (2) untreated ovariectomized rats (OVX), (3) OVX rats treated with OST, (4) OVX rats treated with RAL, (5) OVX rats treated with OST and RAL. Both compounds were administered in the diet. The average dose received was 0.6 ± 0.1 mg for OST and 11.1 ± 1.2 mg for RAL per kg body weight/day. After thirteen weeks, rat activity, muscle weight, structure, gene expression, and serum markers were analyzed. RESULTS: OST increased muscle weight, capillary ratio, insulin-like growth factor 1 (Igf-1) expression, serum phosphorus, uterine weight. RAL decreased muscle weight, capillary ratio, food intake, serum calcium and increased Igf-1 and Myostatin expression, serum follicle stimulating hormone (FSH). OST + RAL increased muscle nucleus ratio, uterine weight, serum phosphorus, FSH and luteinizing hormone and decreased body and muscle weight, serum calcium. Neither treatment changed muscle fiber size. OVX increased body and muscle weight, decreased uterine weight, serum calcium and magnesium. CONCLUSION: OST had beneficial effects on muscle in OVX rats. Side effects of OST on the uterus and serum electrolytes should be considered before using it for therapeutic purposes. RAL and RAL + OST had less effect on muscle and showed endocrinological side effects on pituitary-gonadal axis.

Changes of the subchondral bone microchannel network in early osteoarthritis.

OBJECTIVE: We have identified a 3D network of subchondral microchannels that connects the deep zone of cartilage to the bone marrow (i.e., cartilage-bone marrow microchannel connectors; CMMC). However, the pathological significance of CMMC is largely unknown. Here, we quantitatively evaluated how the CMMC microarchitecture is related to cartilage condition, as well as regional differences in early idiopathic osteoarthritis (OA). METHODS: Two groups of cadaveric female human femoral heads (intact cartilage vs early cartilage lesions) were identified, and a biopsy-based high-resolution micro-CT imaging was employed. Subchondral bone (SB) thickness, CMMC number, maximum and minimum CMMC size, and the CMMC morphology were quantified and compared between the two groups. The effect of joint's region and cartilage condition was examined on each dependent variable. RESULTS: The CMMC number and morphology were affected by region of the joint, but not by cartilage condition. On the other hand, the minimum and maximum CMMC size was changed by both the location on the joint, as well as the cartilage condition. The smallest CMMC were consistently detected at the load-bearing region (LBR) of the joint. Compared to non-pathological subjects, the size of the microchannels was enlarged in early OA, most noticeably at the non-load-bearing region (NLBR) and the peripheral rim (PR) of the femoral head. Furthermore, subchondral bone thinning was observed in early OA as a localized occurrence linked with areas of partial chondral defect. CONCLUSION: Our data point to an enlargement of the SB microchannel network, and a collective structural deterioration of SB in early idiopathic OA.

A combined treatment with selective androgen and estrogen receptor modulators prevents bone loss in orchiectomized rats.

PURPOSE: Enobosarm (EN), a selective androgen receptor modulator and raloxifene (RAL), a selective estrogen receptor modulator, have been shown to improve bone tissue in osteoporotic males. The present study evaluated the effects of a combination therapy of EN and RAL on bone properties in orchiectomized rats compared to the respective single treatments. METHODS: Eight-month-old male Sprague-Dawley rats were either left intact (Non-Orx) or orchiectomized (Orx). The Orx rats were divided into four groups (n = 15 each): 1) Orx, 2) EN treatment (Orx + EN), 3) RAL treatment (Orx + RAL), 4) combined treatment (Orx + EN + RAL). EN and RAL (0.4 mg and 7 mg/kg body weight/day) were applied immediately after Orx with a soy-free pelleted diet for up to 18 weeks. The lumbar spine and femora were examined by micro-CT, biomechanical, histomorphological, ashing, and gene expression analyses. RESULTS: EN exhibited an anabolic effect on bone, improving some of its parameters in Orx rats, but did not affect biomechanical properties. RAL exhibited antiresorptive activity, maintaining the biomechanical and trabecular parameters of Orx rats at the levels of Non-Orx rats. EN + RAL exerted a stronger effect than the single treatments, improving most of the bone parameters. Liver weight increased after all treatments; the kidney, prostate, and levator ani muscle weights increased after EN and EN + RAL treatments. BW was reduced due to a decreased food intake in the Orx + RAL group and due a reduced visceral fat weight in the Orx + EN + RAL group. CONCLUSION: The EN + RAL treatment appeared to be promising in preventing male osteoporosis, but given the observed side effects on liver, kidney, and prostate weights, it requires further investigation.

Combination of selective androgen and estrogen receptor modulators in orchiectomized rats.

PURPOSE: Selective androgen and estrogen receptor modulators, ostarine (OST) and raloxifen (RAL), reportedly improve muscle tissue and offer therapeutic approaches to muscle maintenance in the elderly. The present study evaluated the effects of OST and RAL and their combination on musculoskeletal tissue in orchiectomized rats. METHODS: Eight-month-old Sprague Dawley rats were analyzed. Experiment I: (1) Untreated non-orchiectomized rats (Non-ORX), (2) untreated orchiectomized rats (ORX), (3) ORX rats treated with OST during weeks 0-18 (OST-P), (4) ORX rats treated with OST during weeks 12-18 (OST-T). Experiment II: 1) Non-ORX, (2) ORX, 3) OST-P, (4) ORX rats treated with RAL, during weeks 0-18 (RAL-P), 5) ORX rats treated with OST + RAL, weeks 0-18 (OST + RAL-P). The average daily doses of OST and RAL were 0.4 and 7 mg/kg body weight (BW). Weight, fiber size, and capillarization of muscles, gene expression, serum markers and the lumbar vertebral body were analyzed. RESULTS: OST-P exerted favorable effects on muscle weight, expression of myostatin and insulin growth factor-1, but increased prostate weight. OST-T partially improved muscle parameters, showing less effect on the prostate. RAL-P did not show anabolic effects on muscles but improved body constitution by reducing abdominal area, food intake, and BW. OST + RAL-P had an anabolic impact on muscle, reduced androgenic effect on the prostate, and normalized food intake. OST and RAL improved osteoporotic bone. CONCLUSIONS: The OST + RAL treatment appeared to be a promising option in the treatment of androgen-deficient conditions and showed fewer side effects than the respective single treatments.

Effect of Strontium Ranelate on the Muscle and Vertebrae of Ovariectomized Rats.

Osteoporosis is often accompanied by sarcopenia. The effect of strontium ranelate (SR) on muscle tissue has not been investigated sufficiently. In this study, the effect of different SR treatments on muscle was studied. Additionally, the lumbar vertebrae were analyzed. Three-month-old female rats were divided into five groups (n = 12): Group 1: untreated (NON-OVX); Group 2: ovariectomized and left untreated (OVX); Group 3: SR after OVX until the study ended (13 weeks, SR prophylaxis and therapy = pr+th); Group 4: OVX and SR for 8 weeks (SR prophylaxis = pr); Group 5: SR for 5 weeks from the 8 week after OVX (SR therapy = SR th). SR was applied in food (630 mg/kg body weight). The size of muscle fibers, capillary density, metabolic enzymes, and mRNA expression were assessed in soleus, gastrocnemius, and longissimus muscles. The vertebral bodies underwent micro-CT, biomechanical, and ashing analyses. In general, SR did not alter the muscle histological parameters. The changes in fiber size and capillary ratio were related to the body weight. Myostatin mRNA was decreased in Sr pr+th; protein expression was not changed. SR th led to increase in mRNA expression of vascular endothelial growth factor (Vegf-B). In lumbar spine, SR pr+th enhanced biomechanical properties, bone mineral density, trabecular area, density, and thickness and cortical density. The reduced calcium/phosphate ratio in the SR pr+th group indicates the replacement of calcium by strontium ions. SR has no adverse effects on muscle tissue and it shows a favorable time-dependent effect on vertebrae. A functional analysis of muscles could verify these findings.

Real-life experience with first generation HCV protease inhibitor therapy in Germany: The prospective, non-interventional PAN cohort.

BACKGROUND AND AIMS: The efficacy and safety of peginterferon alfa-2a (PEG-IFN) plus ribavirin (RBV) and either boceprevir (BOC) or telaprevir (TVR), and physician adherence to treatment algorithms were evaluated in patients included in an ongoing non-interventional study (PAN) enrolling adults with chronic hepatitis C virus (HCV) infection managed in German office-based practices. METHODS: The analysis included HCV genotype 1-infected, treatment-naïve and treatment-experienced patients treated with BOC or TVR. Demographic, treatment history, virological response, safety, and patient management data were collected. RESULTS: Of a total 1087 patients, 58.1 % achieved sustained virological responses (SVR). Response rates were higher in treatment-naïve (BOC 55 %; TVR 63.4 %) and prior relapse patients (BOC 63.2 %; TVR 74.5 %) versus previous null-responders (BOC 14.3 %; TVR 25 %). The most commonly reported adverse event overall was fatigue (60.6 %); 45.8 % patients experienced hemoglobin < 10 g/dL. Patients with cirrhosis had lower rates of SVR versus those without (42.9 % vs. 60.7 %, respectively), and had a higher incidence of serious adverse events (SAEs) (16.7 % vs. 8.6 %, respectively) and treatment discontinuation (44.6 % vs. 25.2 %, respectively). According to recommended response-guided treatment algorithms, about 70 % of patients were managed appropriately, 11/10 % (BOC/TVR) received unnecessarily extended therapy, and 19/7 % (BOC/TVR) received inappropriately shortened therapy. CONCLUSIONS: The efficacy and safety of BOC- and TVR-based triple therapy in this large, "real-world" cohort were largely comparable to that reported in pivotal clinical trials, although SVR rates were lower overall. Recommended futility or treatment extension rules were violated in a substantial proportion of patients with potential implications for response, adverse events and costs.

Treatment of osteoporosis using a selective androgen receptor modulator ostarine in an orchiectomized rat model.

PURPOSE: The selective androgen receptor modulator ostarine has been shown to have advantageous effects on skeletal tissue properties, reducing muscle wasting and improving physical function in males. However, data on effects in male osteoporosis remain limited. In this study, the effects of ostarine on osteoporotic bone were evaluated in a rat model of male osteoporosis and compared with those of testosterone treatments. METHODS: Eight-month-old male Sprague-Dawley rats were either non-orchiectomized to serve as a healthy control (Non-Orx, Group 1) or orchiectomized (Orx, Groups 2-6) and then grouped (n = 15/group): (1) Non-Orx, (2) Orx, (3) Ostarine Therapy, (4) Testosterone Therapy, (5) Ostarine Prophylaxis and (6) Testosterone Prophylaxis. Prophylaxis treatments started directly after orchiectomy and continued for 18 weeks, whereas Therapy treatments were initiated 12 weeks after Orx. Ostarine and Testosterone were applied orally at daily doses of 0.4 and 50 mg/kg body weight, respectively. The lumbar vertebral bodies and femora were analyzed using biomechanical, micro-CT, ashing, and gene expression analyses. RESULTS: Ostarine Prophylaxis showed positive effects in preventing osteoporotic changes in cortical and trabecular bone (femoral trabecular density: 26.01 ± 9.1% vs. 20.75 ± 1.2% in Orx and in L4: 16.3 ± 7.3% vs 11.8 ± 2.9% in Orx); biomechanical parameters were not affected; prostate weight was increased (0.62 ± 0.13 g vs 0.18 ± 0.07 g in Orx). Ostarine Therapy increased solely the cortical density of the femur (1.25 ± 0.03 g/cm3 vs. 1.18 ± 0.04 g/cm3 in Orx); other bone parameters remained unaffected. Testosteron Prophylaxis positively influenced cortical density in femur (1.24 ± 0.05 g/cm3 vs. 1.18 ± 0.04 g/cm3 in Orx); Test. Therapy did not change any bony parameters. CONCLUSION: Ostarine Prophylaxis could be further investigated as a preventative treatment for male osteoporosis, but an androgenic effect on the prostate should be taken into consideration, and combination therapies with other anti-osteoporosis agents could be considered.

Predicting the disaster - The role of CRP in acetabular surgery.

OBJECTIVES: Acetabular fractures represent a complex surgical challenge. Given the heterogenous fracture pattern, the patient characteristics and spectrum of complications demand individual solutions. Surgical site infections (SSI) threaten osteosynthesis, and early detection of them and treatment remain crucial. What is the value of postoperative C-reactive protein (CRP) in this group of patients as well as its normal course? DESIGN & METHODS: 115 patients with isolated fractures of the acetabulum were retrospectively evaluated. CRP, white blood cell count (WBC) and fracture patterns as well as patient characteristics were assessed for 20 days following operative fixation of the acetabular fracture (n = 71) and in fractures that were managed conservatively (n = 44). RESULTS: Twelve patients suffered an infectious complication. With a one-phase decay, 70.55% of the variance of postoperative CRP kinetics was predicted. To anticipate maximum CRP as well as an infection, the preoperative CRP represented the best prognostic parameter. To predict an infection, the single variable "peak CRP value above 100 mg/l" resulted in a sensitivity and specificity of 91.67% and 36.21%, respectively. Combining a second peak of CRP with maximum CRP and day 5 CRP value for receiver-operating characteristic (ROC) analysis resulted in 83.3% and 88.1%, respectively. CONCLUSIONS: Predicting surgical site infections after an acetabular fracture is most predictive when analyzing the maximum overall CRP, the second peak and the CRP after day 5. With a combination of these parameters, a sensitivity and specificity of 83.3% and 88.1% to detect an infection was achieved.

Distinct roles of free leptin, bound leptin and soluble leptin receptor during the metabolic-inflammatory response in patients with liver cirrhosis.

BACKGROUND: Alteration of the leptin system appears to play a role in the inflammatory-metabolic response in catabolic diseases such as chronic liver diseases. AIM: To investigate the association between leptin components, inflammatory markers and hepatic energy and substrate metabolism. METHODS: We investigated in vivo hepatic substrate and leptin metabolism in 40 patients employing a combination of arterial and hepatic vein catheterization techniques and hepatic blood flow measurements. In addition to metabolic, inflammatory and neuroendocrine parameters, circulating levels of free leptin, bound leptin and soluble leptin receptor were determined. RESULTS: Compared with controls, bound leptin and soluble leptin receptor levels were significantly elevated in cirrhosis, while free leptin did not increase. In cirrhosis bound leptin was correlated with soluble leptin receptor (r = 0.70, P < 0.001). Free leptin was positively correlated with metabolic parameters such as energy storage (body fat mass; r = 0.36, P < 0.05), insulin and insulin resistance (r = 0.48; r = 0.46, P < 0.01) as well as with hepatic glucose and energy release (r = 0.35 and r = 0.40, P < 0.05). In contrast, bound leptin and soluble leptin receptor were linked to proinflammatory cytokines and sympathetic activity (r = 0.61 and r = 0.56, P < 0.01). CONCLUSION: The components of the leptin system (free leptin, bound leptin and soluble leptin receptor) have distinct roles in metabolic and inflammatory processes in patients with liver cirrhosis. The better understanding of this metabolic and inflammatory tissue-repair response may lead to innovative new therapeutic strategies in liver disease as well as in various other catabolic diseases.

Real-world use, effectiveness and safety of anti-viral treatment in chronic hepatitis C genotype 3 infection.

BACKGROUND: Treatment of chronic hepatitis C genotype 3 (GT3) is more challenging compared with other genotypes. Since 2014, several new treatment regimens have been approved but sometimes based on limited data. AIM: To validate the use, effectiveness and safety of anti-viral treatment in chronic hepatitis C genotype 3 infection under real-word conditions. METHODS: The German Hepatitis C-Registry is a large national non-interventional real-world study for patients with chronic hepatitis C. A total of 1322 GT3 patients were enrolled (211 untreated and 1111 treated patients). RESULTS: Between February 2014 and September 2015, five different treatment strategies have been used (PegIFN+RBV, PegIFN+RBV+SOF, SOF+RBV, DCV+SOF±RBV, LDV/SOF±RBV). Treatment uptake and use of treatment concepts changed markedly and rapidly during the study influenced by new approvals, guideline recommendations, and label updates. PegIFN-based therapies constantly declined while DCV-based therapies increased with one interruption after the approval of LDV/SOF, which was frequently used until new guidelines recommended not using this combination for GT3. Per-protocol SVR ranged from 80.9% in the PegIFN+RBV group to 96.1% in PegIFN+RBV+SOF treated patients. Treatment-experienced patients with cirrhosis showed a suboptimal SVR of 68% for SOF+RBV but a high SVR of 90-95% for DCV+SOF±RBV. The safety analysis showed more adverse events and a stronger decline of haemoglobin for RBV containing regimens. CONCLUSIONS: Real-world data can validate the effectiveness and safety for treatment regimens that had previously been approved with limited data, in particular for specific subgroups of patients. The present study demonstrates how rapid new scientific data, new treatment guidelines, new drug approvals and label changes are implemented into routine clinical practice today.

Hypermetabolism in clinically stable patients with liver cirrhosis.

BACKGROUND: Hypermetabolism has a negative effect on prognosis in patients with liver cirrhosis. Its exact prevalence and associations with clinical data, the nutritional state, and beta-adrenergic activity are unclear. OBJECTIVE: We investigated resting energy expenditure (REE) in 473 patients with biopsy-proven liver cirrhosis. DESIGN: This was a cross-sectional study with a controlled intervention (beta-blockade) in a subgroup of patients. RESULTS: Mean REE was 7.12 +/- 1.34 MJ/d and correlated closely with predicted values (r = 0.70, P < 0.0001). Hypermetabolism was seen in 160 patients with cirrhosis (33.8% of the study population). REE was > 30% above the predicted value in 41% of the hypermetabolic patients with cirrhosis. Hypermetabolism had no association with clinical or biochemical data on liver function. REE correlated with total body potassium content (TBP; r = 0.49, P < 0.0001). Hypermetabolic patients had lower than normal body weight and TBP (P < 0.05). About 47% of the variance in REE could be explained by body composition whereas clinical state could maximally explain 3%. Plasma epinephrine and norepinephrine concentrations were elevated in hypermetabolic cirrhotic patients (by 56% and 41%, respectively; P < 0.001 and 0.01). Differences in REE from predicted values were positively correlated with epinephrine concentration (r = 0.462, P < 0.001). Propranolol infusion resulted in a decrease in energy expenditure (by 5 +/- 3%; P < 0.05), heart rate (by 13 +/- 4%; P < 0.01), and plasma lactate concentrations (by 32 +/- 12%; P < 0.01); these effects were more pronounced in hypermetabolic patients (by 50%, 33%, and 68%, respectively; each P < 0.05). CONCLUSIONS: Hypermetabolism has no association with clinical data and thus is an extrahepatic manifestation of liver disease. Increased beta-adrenergic activity may explain approximately 25% of hypermetabolism.

Prostaglandin E plus famciclovir--a new concept for the treatment of severe hepatitis B after liver transplantation.

New concepts for the treatment of hepatitis B in immunocompromised patients are urgently needed. We describe our first experience with the new antiviral agent famciclovir in combination with a short course of prostaglandin E in a patient with severe hepatitis B after liver transplantation. Initial treatment with prostaglandin E reduced the inflammatory activity, as measured by transaminase activities, but did not affect viral replication. Consecutive long-term treatment with famciclovir further normalized liver function and profoundly suppressed viral replication. HBeAg and HBV-DNA -PCR all became negative and only HBsAg persisted. Histology documented marked reduction of cellular infiltration. The patient completely recovered and is back to regular work as a teacher.

Development, stability, and clinical correlations of allogeneic microchimerism after solid organ transplantation.

To assess the development, stability, and clinical relevance of donor-type microchimerism, skin and blood were analyzed in heart (n = 53) and liver (n = 18) transplant recipients by nested polymerase chain reaction. Microchimerism was detectable in 40 (75%) and 13 (72%) patients after heart and liver transplantation, respectively. In heart transplantation, chimerism-positive patients showed a lower frequency of acute rejection as compared with negative patients, although this was only of borderline statistical significance. Repeated intraindividual analyses demonstrated variable patterns of microchimerism over time, but changes did not correlate to the clinical state. In liver transplantation, chimeric state showed no clear correlation with the patients' immunological situation. Our results demonstrate that peripheral microchimerism frequently develops after different types of organ transplantation and represents a dynamic process but without diagnostic value to predict the immunological risk for individual patients.

Clinical and subclinical acute rejection early after liver transplantation: contributing factors and relevance for the long-term course.

Routine transplant aspiration cytology (TAC) after liver transplantation gives detailed information that concerns immunologic events in the graft. TAC can be helpful for diagnosis of acute rejection, but it also detects morphological signs of rejection without clinical correlate ("subclinical rejection"). The aim of this study was to systematically evaluate factors that influence the development of early clinical and subclinical rejection and to analyze the relevance of these early immunologic processes for the long-term course. The study includes the course of 340 patients after liver transplantation between 1988 and 1995 in whom TAC was performed routinely and who were followed for a minimum of 3 years. TAC findings were correlated with the following various clinical parameters: (1) Overall early clinical rejection occurred in 17.4%, subclinical rejection in 59.1%, and no immune activation was seen in 23.5% of patients. (2) Incidence of early clinical and subclinical rejection was markedly influenced by type of immunosuppression. (3) Basic disease and extent of preservation injury had only a minor influence; there was a trend towards lower early rejection associated with more severe preservation damage, increased patient age, and early retransplantation. (4) Presence of early clinical or subclinical rejection was not associated with a higher incidence of chronic dysfunction. (5) Falsely indicated antirejection treatment was associated with inferior graft survival. Subclinical rejection is very frequent early after liver transplantation, requires no treatment, and has no long-term adverse effect. Incidence of early clinical rejection is mainly determined by initial immunosuppression; its occurrence has no negative long-term effects and may even be associated with a lower risk for later immunological complications. Thus, the incidence of early acute rejection is no adequate parameter for evaluating the quality of an immunosuppressive treatment protocol.

Improvement of acute and chronic renal dysfunction in liver transplant patients after substitution of calcineurin inhibitors by mycophenolate mofetil.

BACKGROUND: Renal dysfunction caused by treatment with the calcineurin inhibitors (CNI) is a major problem in the long-term course after liver transplantation. PATIENTS: In 22 liver graft recipients with renal dysfunction and stable graft function between 3 weeks and 12 years after transplantation, CNI were substituted by MMF at a final dose of 1.5-3 g/day between October 1996 and October 1998. METHODS: In a prospective non-randomized study, the development of renal function, the side effects of MMF medication, and the stability of liver function were analyzed for a mean follow-up of 15 months. Results. (1) MMF was withdrawn in four patients for major side effects between 1 and 7 months after study entry; eight patients had minor side effects. (2) Six months after study entry, renal function had improved in 17 of the 22 study patients; mean serum creatinine +/-SD (micromol/L) was 201+/-77 at entry and 153+/-65 after 3 months (P<0.001). (3) Improvement occurred in 11 of 15 patients with creatinine elevation > or =12 months and in 6 of 6 patients with creatinine elevation < or =6 months. (4) One patient developed transient liver dysfunction and a second required retransplantation for progressive cholestasis but without signs of rejection. CONCLUSIONS: In patients who undergo liver transplantation, substitution of CNI by MMF leads to improvement of acute as well as chronic renal dysfunction in most cases. Side effects of MMF may be limiting in some patients, and the immunological consequences remain to be studied.

Famciclovir treatment of chronic hepatitis B in heart transplant recipients: a prospective trial.

UNLABELLED: Hepatitis B may take a rapid and aggressive course in patients under immunosuppression. Nucleoside analogues have been shown to suppress viral replication effectively. To investigate the effect of famciclovir in immunosuppressed patients, 21 heart transplant recipients with chronic hepatitis B infection were included in a prospective study. PATIENTS AND METHODS: Patients have been treated with Famciclovir for a median of 14 months. Hepatitis B virus replication and biochemical parameters were regularly tested and liver biopsies were taken before treatment and after a median time of 7 months. HBV-polymerase was sequenced in all patients before therapy and in those patients who experienced virological breakthrough. RESULTS: Nineteen patients were treated for at least 6 months. Hepatitis B virus-DNA levels declined in all patients and became negative in 8 patients. Mean hepatitis B virus-DNA levels decreased from 199+/-269 to 34+/-53 pg/ml after 24 weeks (P=0.003). During treatment HBeAg became negative in five patients. Mean alanine aminotransferase decreased from 42+/-26 to 24+/-10 U/L (P=0.006). Histological analysis revealed improved inflammatory activity according to the Ishak-score in 11/16 (69%) patients. Total inflammatory activity scores decreased from 8 to 6 (median, NS), but interface hepatitis score (P=0.02) and lobular inflammation score (P=0.006) improved significantly. Median fibrosis scores fell from 5 to 3 (P=0.002). Three patients developed virological breakthrough on famciclovir after 7, 8, and 26 months of treatment showing HBV-polymerase amino acid changes L528 M, S567A, and I581K, respectively. CONCLUSIONS: Famciclovir improves not only biochemical and virological features but also hepatic inflammation and liver fibrosis in patients with chronic hepatitis B under heavy immunosuppression. Virological breakthrough may develop and requires close monitoring.

Comparison of matrix metalloproteinase expression in normal and cirrhotic human liver.

To study the extend of ongoing tissue remodelling in end-stage cirrhosis, the expression of different matrix metalloproteinases [interstitial collagenase (MMP-1), Mr 72000 gelatinase (MMP-2), stromelysin-1 (MMP-3) and stromelysin-3 (MMP-11)] and of TIMP-1 was studied in 13 cirrhotic livers explanted at transplantation. The results were compared with those obtained in normal liver. Western blot, northern blot, ELISA, RT-PCR and zymogram analysis were used. Proenzymes of stromelysin-1 and -3, interstitial collagenase and Mr 72000 gelatinase were positive in normal liver, while activated enzymes were not detectable in western blot analysis. In cirrhosis proenzyme levels of the studied MMPs were reduced to a mean of 60-70%, but mRNA expression and gelatin-degrading activity increased. TIMP-1 expression was detectable on mRNA level and by ELISA in normal liver and also increased in cirrhosis. Our results show that mRNA expression of certain matrix metalloproteinases is increased in end-stage liver cirrhosis, while the amount of proenzyme is decreased, indicating enhanced MMP proenzyme turnover. These data suggest that besides increased TIMP-1 activity, altered MMP expression may also play a part in fibroproliferation in liver disease.

Tacrolimus (FK 506) biotransformation in primary rat hepatocytes depends on extracellular matrix geometry.

Established in vitro models for studies of hepatic drug biotransformation include the use of primary hepatocytes. In normal liver the space of Disse provides the possibility of bilateral attachment to extracellular matrix for each hepatocyte. This configuration is disrupted by the cell isolation procedure of normal liver tissue, which delivers suspensions of round shaped cells. In standard culture configurations this unphysiologic cell shape terminates in a morphological dedifferentiation and inability to biotransform drugs. This study analyses the relevance of extracellular matrix geometry in hepatocyte monolayer configurations for expression and activity of cytochrome P450 3A. This enzyme is involved in the biotransformation of a large number of pharmaceuticals including the immunosuppressants tacrolimus and sirolimus. Morphological analysis of primary rat hepatocytes cultured with and without overlay of collagen type I was performed by transmission and scanning electron microscopy. Expression and activity of cytochrome P450 3A was studied by Western blot and the use of two model drugs specific for this enzyme. To this purpose the immunosuppressive drugs tacrolimus and sirolimus were used. Metabolites were analyzed by HPLC and HPLC/MS. Two sided attachment to extracellular matrix induces profound changes of the hepatocellular morphology in vitro resulting in the reconstitution of a polyhedric cell shape. This phenomenon is paralleled by an enhanced expression of cytochrome P450 3A and corresponding metabolic activity. As shown for tacrolimus biotransformation, the model may be useful to study complex metabolic patterns. In addition this model may facilitate studies of the kinetics of hepatocellular drug biotransformation in a setting with prolonged stability.

Are patients with liver cirrhosis hypermetabolic?

Hypermetabolism is not a constant feature of liver cirrhosis. It may occur in up to 18% of cirrhotics. Most of the deviations are due to increases in resting energy expenditure (REE). Dietary induced thermogenesis (DIT) is normal or slightly increased whilst the thermic effect of exercise TEE is of minor importance in cirrhosis. The increase in REE which reflects a systemic manifestation of liver disease cannot be identified by the clinical and biochemical measures of liver function. An increased REE is frequently seen in malnourished patients and this is mainly due to disproportional loss in muscle mass. Some cirrhotic patients cannot reduce REE in response to weight loss. This problem is not specific for liver cirrhosis but is also seen in other cachectic groups of patients. Adjustment of REE per kg fat free mass (FFM) may lead to erroneous conclusions (i) because of the non linearity of REE over the range of FFM and (ii) the different contributions of muscle mass and non-muscle body cell mass (BCM) to FFM over the range of FFM. There is circumstantial evidence that the metabolic rate per kg BCM is increased in malnourished cirrhotics. More specifically, cirrhosis increases in REE are associated with a deterioration in hepatic circulation. Increased sympathetic nervous system activity is frequently seen in cirrhosis and may provide a link between between reduced nutritive portal flow and increased whole body oxygen consumption. Increased REE is also associated with weight loss, a poorer liver function and a higher mortality after liver transplantation and thus may have prognostic value. Taken together, REE is variable in patients with cirrhosis. Hypermetabolism is seen in malnourished patients and those with impaired splanchnic hemodynamics. Hypermetabolism is associated with a poorer outcome after liver transplantation.

An organotypical in vitro model of the liver parenchyma for uptake studies of diagnostic MR receptor agents.

Testing of receptor-specific MR contrast agents targeted to the liver is hampered by a shortage of viable in vitro models with in vivo-like hepatocellular morphology. Coated pits are ultrastructural signs of an active receptor mediated endocytosis in hepatocytes. Expression of coated pits by matrix overlaid hepatocytes was studied by transmission electron microscopy. Binding of a rhodaminated asialoglycoprotein receptor agent (MION-ASF-rh) was assessed by fluorescence microscopy. Fluorescence of cells exposed to MION-ASF-rh with D(+)-galactose reduced fluorescent light emission to a level of 58% of MION-ASF-rh-induced fluorescence. After preincubation with the hepatotoxin CCl4 a dose-dependent decrease in fluorescent light emission resulted. Hepatocytes maintained a homogeneous cell surface expression, with microprojections, coated pits, and vesicles on both sinusoidal surfaces. Matrix overlaid primary hepatocytes constitute a viable, morphologically and functionally differentiated model. This model can be used to study receptor binding, uptake, and blockage of diagnostic magnetopharmaceuticals under controlled conditions.