Improvement in liver fibrosis, functionality and hemodynamics in CCI4-cirrhotic rats after injection of the Liver Growth Factor.

BACKGROUND/AIMS: Most substances used in experimental models of cirrhosis are chosen either as protectors of lipid peroxidation, as antifibrogenic agents or as vitamins, among others. In this report, we analyze the improvement produced, in established cirrhosis (CCl4 plus phenobarbital) in rats, by intraperitoneal injection of Liver Growth Factor, a hepatic mitogen with activity both in vivo and in vitro. METHODS: Following confirmation of CCl4-induced cirrhosis, Liver Growth Factor (4.5 microg per ratx2 injections/week for 3 weeks) was administered to one group of rats (Cirr+LGF). The remaining rats (Cirr) received saline. The groups were compared in terms of serum enzymes, tissue damage, total liver collagen, collagenase activity, microsomal enzyme activities, splanchnic and systemic hemodynamics and portosystemic shunting. RESULTS: Treatment of rats presenting CCl4-induced cirrhosis with Liver Growth Factor decreased serum aminotransferase levels and increased levels of serum albumin and total protein. The Liver collagen content was lower in rats treated with Liver Growth Factor (2.96 vs. 4.32 mg/g liver, p<0.01). Microscopic studies revealed that the livers of rats receiving Liver Growth Factor showed decreases in fibrosis, necrosis and inflammatory infiltration, as well as a recovery of architectural integrity. Liver function was improved after treatment with Liver Growth Factor, as indicated by the rate constant for elimination of aminopyrine, which increased from 0.0063 to 0.0170 (p<0.05). This increase was accompanied by a higher total amount of cytochrome P-450 as well as of certain P-450 isoenzymes, especially those that are hormone-dependent, such as P-450 3A. The improved liver histology and function observed in Cirr+LGF rats was associated with decreases in portal pressure (14.4 vs. 9.4 mm Hg, p<0.01) and portosystemic shunting (55.8 vs. 11.5%, p<0.01), as well as increases in mean arterial pressure and systemic vascular resistance, and a reduction in ascites. CONCLUSIONS: Administration of the hepatic mitogen, Liver Growth Factor, to CCl4-cirrhotic rats decreased liver collagen and reorganized the hepatic extracellular matrix, resulting in an improvement in liver function, reduced portal pressure and amelioration of ascites.     

Shift from biliary to urinary elimination of acetaminophen-glucuronide in acetaminophen-pretreated rats

Despite its toxicity, acetaminophen (APAP) is used increasingly as an analgesic, antipyretic, and anti-inflammatory agent. We examined the effect of prior exposure to APAP on its biliary and urinary elimination. The biliary and urinary elimination of a test dose of APAP (150 mg/kg i.v.) was determined in male Wistar rats 24 h after pretreatment with vehicle, a single dose (1.0 g/kg i.p.), or increasing daily doses (0.2, 0.3, 0.6, and 1.0 g/kg/day i.p.) of APAP. Although elimination of the parent APAP was minimally affected, biliary excretion of APAP glucuronide was significantly decreased 70 and 80%, whereas urinary excretion was significantly increased 90 and 100% in the groups pretreated with single and repeated doses of APAP, respectively, relative to vehicle controls. Western analysis and confocal immunofluorescent microscopy indicated a marked increase in hepatic expression of multidrug resistance-associated protein 3 (Mrp3) in both groups pretreated with APAP, relative to expression of Mrp2. ATP-dependent transport of [3H]taurocholate, an Mrp3 substrate, was significantly increased in basolateral liver plasma membrane vesicles from rats pretreated with repeated doses of APAP relative to controls. Enterohepatic recirculation of APAP glucuronide after administration of the same test dose of the drug was significantly decreased in rats pretreated with repeated doses of APAP. These data indicate that APAP pretreatment induced a shift from biliary to urinary elimination of APAP glucuronide, consistent with the increased expression of Mrp3 in the basolateral domain of the hepatocyte. We postulate that decreased enterohepatic recirculation contributes to decreased APAP hepatotoxicity by reducing liver exposure.     

Azathioprine acts upon rat hepatocyte mitochondria and stress-activated protein kinases leading to necrosis: protective role of N-acetyl-L-cysteine

Azathioprine is an immunosuppressant drug widely used. Our purpose was to 1) determine whether its associated hepatotoxicity could be attributable to the induction of a necrotic or apoptotic effect in hepatocytes, and 2) elucidate the mechanism involved. To evaluate cellular responses to azathioprine, we used primary culture of isolated rat hepatocytes. Cell metabolic activity, reduced glutathione, cell proliferation, and lactate dehydrogenase release were assessed. Mitochondria were isolated from rat livers, and swelling and oxygen consumption were measured. Mitogen-activated protein kinase pathways and proteins implicated in cell death were analyzed. Azathioprine decreased the viability of hepatocytes and induced the following events: intracellular reduced glutathione (GSH) depletion, metabolic activity reduction, and lactate dehydrogenase release. However, the cell death was not accompanied by DNA laddering, procaspase-3 cleavage, and cytochrome c release. The negative effects of azathioprine on the viability of hepatocytes were prevented by cotreatment with N-acetyl-L-cysteine. In contrast, 6-mercaptopurine showed no effects on GSH content and metabolic activity. Azathioprine effect on hepatocytes was associated with swelling and increased oxygen consumption of intact isolated rat liver mitochondria. Both effects were cyclosporine A-sensitive, suggesting an involvement of the mitochondrial permeability transition pore in the response to azathioprine. In addition, the drug's effects on hepatocyte viability were partially abrogated by c-Jun N-terminal kinase and p38 kinase inhibitors. In conclusion, our findings suggest that azathioprine effects correlate to mitochondrial dysfunction and activation of stress-activated protein kinase pathways leading to necrotic cell death. These negative effects of the drug could be prevented by coincubation with N-acetyl-L-cysteine.     

Severe Serotonin Depletion after Conditional Deletion of the Vesicular Monoamine Transporter 2 Gene in Serotonin Neurons: Neural and Behavioral Consequences

The vesicular monoamine transporter type 2 gene (VMAT2) has a crucial role in the storage and synaptic release of all monoamines, including serotonin (5-HT). To evaluate the specific role of VMAT2 in 5-HT neurons, we produced a conditional ablation of VMAT2 under control of the serotonin transporter (slc6a4) promoter. VMAT2^sert−cre mice showed a major (−95%) depletion of 5-HT levels in the brain with no major alterations in other monoamines. Raphe neurons contained no 5-HT immunoreactivity in VMAT2^sert−cre mice but developed normal innervations, as assessed by both tryptophan hydroxylase 2 and 5-HT transporter labeling. Increased 5-HT_1A autoreceptor coupling to G protein, as assessed with agonist-stimulated [³⁵S]GTP-γ-S binding, was observed in the raphe area, indicating an adaptive change to reduced 5-HT transmission. Behavioral evaluation in adult VMAT2^sert−cre mice showed an increase in escape-like reactions in response to tail suspension and anxiolytic-like response in the novelty-suppressed feeding test. In an aversive ultrasound-induced defense paradigm, VMAT2^sert−cre mice displayed a major increase in escape-like behaviors. Wild-type-like defense phenotype could be rescued by replenishing intracellular 5-HT stores with chronic pargyline (a monoamine oxidase inhibitor) treatment. Pargyline also allowed some form of 5-HT release, although in reduced amounts, in synaptosomes from VMAT2^sert−cre mouse brain. These findings are coherent with the notion that 5-HT has an important role in anxiety, and provide new insights into the role of endogenous 5-HT in defense behaviors.     

Activation of the immune response against Infectious Bursal Disease Virus after intramuscular inoculation of an intermediate strain

Infectious bursal disesase is a highly contagious, wide spread immunosuppressive chicken disease caused by the Infectious Bursal Disease Virus (IBDV). IBDV is a two segmented double-strand RNA virus, member of the Birnaviridae family. In order to study the interaction between IBDV and the immune system, chickens were exposed to an intermediate IBDV strain by intramuscular route, and using Real Time PCR the expression of a panel of avian cytokines and chemokines in duodenum, spleen and bursa of Fabricius was analyzed. Also, splenic nitrite (NO₂) production and the frequencies of different mononuclear cell populations were evaluated by Griess reaction and flow cytometry, respectively. Intramuscular (i.m.) IBDV inoculation promoted an over expression of proinflammatory cytokines IL-6, IL-15 and gIFN in spleen, which correlated with an increase of gIFN plasma concentration measured by ELISA, together with an increment of NO₂ concentration in splenocyte supernatants at 1 dpi. Results obtained in the present work showed that IBDV of intermediate virulence, given i.m., induced similar effects to those previously described for highly virulent IBDV in early innate immune responses. Considering that the i.m. route is the route of choice for the delivery of new generation vaccines, and that the use of recombinant antigens also requires the addition of adjuvants for proper immune stimulation, results presented here could contribute to identify suitable cytokines to be used or to be stimulated when utilizing subunit vaccines, for the improvement of prevention tools for avian health.     

MDR1 polymorphisms and response to azathioprine therapy in patients with Crohn's disease

BACKGROUND: To investigate the contribution of multidrug resistance 1 (MDR1) gene pharmacogenetics (G2677T/A and C3435T) to the efficacy of azathioprine in inducing remission in patients with Crohn's disease (CD). METHODS: A cohort of 327 unrelated Spanish patients with CD recruited from a single center was studied. All patients were rigorously followed up for at least 2 years (mean time, 11.5 years). A case-control analysis of MDR1 G2677T/A and C3435T SNPs and 2 loci haplotypes in 112 steroid-dependent CD patients treated with azathioprine was performed. Patients were classified on the basis of response to azathioprine. RESULTS: A total 76 patients treated with azathioprine for longer than 3 months were included. Remission was achieved in 42 CD patients (55.3%). A higher frequency of the 2677TT genotype was found in nonresponders than in responders (17.65% versus 7.14%; OR = 2.8; 95% CI; 0.6-12.1; P = 0.11). Nonresponders to azathioprine were found to have a higher frequency of the 3435TT genotype than did CD patients who had achieved clinical remission (17.64% versus 4.76%; OR = 4.3; 95% CI, 0.8-22.8; P = 0.06). The 2677T/3435T haplotype was also more abundant in nonresponders (29.4% versus 20.2%), whereas the 2677G/3435C haplotype was more frequent in responders (58.3% versus 47.1%). Lack of response to azathioprine therapy in CD patients was 1.8-fold greater in carriers of the 2677T/3435T haplotype than in carriers of the 2677G/3435C haplotype (OR = 1.8; 95% CI, 0.82-3.9; P = 0.14). CONCLUSIONS: The results of our study indicate higher frequencies of the 2677TT and 3435TT genotypes and the 2677T/3435T haplotype in CD patients who did not respond to azathioprine. Additional replications in independent populations would confirm the real impact of these polymorphisms in response to azathioprine therapy.     

Some new food for thought: The role of vitamin D in the mental health of older adults

Vitamin D, a multipurpose steroid hormone vital to health, has been increasingly implicated in the pathology of cognition and mental illness. Hypovitaminosis D is prevalent among older adults, and several studies suggest an association between hypovitaminosis D and basic and executive cognitive functions, depression, bipolar disorder, and schizophrenia. Vitamin D activates receptors on neurons in regions implicated in the regulation of behavior, stimulates neurotrophin release, and protects the brain by buffering antioxidant and anti-inflammatory defenses against vascular injury and improving metabolic and cardiovascular function. Although additional studies are needed to examine the impact of supplementation on cognition and mood disorders, given the known health benefits of vitamin D, we recommend greater supplementation in older adults.     

Bisphosphonate-related osteonecrosis of the jaw: clinical correlations with computerized tomography presentation

The aim of this study was to correlate clinical and computerized tomography (CT) features of bisphosphonate-related osteonecrosis of the jaws (BRONJ). All ONJ patients for whom there was complete CT scan imaging were eligible. Selected clinical parameters retrieved from their medical records were analyzed for correlation with CT parameters. The clinical presentation of BRONJ was supported by findings in CT imaging in 78.3%. The lesion’s size on CT correlated with the presence of purulent secretion (p?=?0.03). When sequestrum was present, the median lesion’s size on CT was relatively big (28 mm, range 21–43 mm). The mandibular canal cortex was never breached. CT has reasonable detection competence for diagnosing BRONJ. Purulent secretion indicates the likelihood that a more extensive involvement will be displayed on CT. A large lesion on CT should raise the index of suspicion for sequestrum. The CT appearance of a continuous cortex of the mandibular canal may serve as a differential parameter between BRONJ and metastasis to the jaw.