pH Dependence of a GPR4 Selective Antagonist Hampers Its Therapeutic Potential.

Inflammatory bowel disease (IBD) is characterized by chronic mucosal inflammation of the gastrointestinal tract and is associated with extracellular acidification of mucosal tissue. Several extracellular pH-sensing receptors, including G protein-coupled receptor 4 (GPR4), play an important role in the regulation of inflammatory and immune responses, and GPR4 deficiency has been shown to be protective in IBD animal models. To confirm the therapeutic potential of GPR4 antagonism in IBD, we tested Compound 13, a selective GPR4 antagonist, in the interleukin 10-/- mouse model of colitis. Despite good exposures and albeit there was a trend toward improvement for a few readouts, Compound 13 treatment did not improve colitis in this model, and there were no signs of target engagement. Interestingly, Compound 13 behaved as an "orthosteric" antagonist, i.e., its potency was pH dependent and mostly inactive at pH levels lower than 6.8 with preferential binding to the inactive conformation of GPR4. Mutagenesis studies confirmed Compound 13 likely binds to the conserved orthosteric binding site in G protein-coupled receptors, where a histidine sits in GPR4 likely preventing Compound 13 binding when protonated in acidic conditions. While the exact mucosal pH in the human disease and relevant IBD mice models is unknown, it is well established that the degree of acidosis is positively correlated with the degree of inflammation, suggesting Compound 13 is not an ideal tool to study the role of GPR4 in moderate to severe inflammatory conditions. SIGNIFICANCE STATEMENT: Compound 13, a reported selective GPR4 antagonist, has been widely used to assess the therapeutic potential of GPR4, a pH-sensing receptor, for numerous indications. Its pH dependence and mechanism of inhibition identified in this study clearly highlights the limitations of this chemotype for target validation.

Hepatocyte growth factor reverses cholemic nephropathy associated with α-naphthylisothiocyanate-induced cholestasis in mice.

Hepatocyte growth factor (HGF) has been proved to protect the liver against α-naphthylisothiocyanate (ANIT)-induced cholestasis by acting as an antioxidant agent and redirecting toxic biliary solutes towards blood for urinary excretion. However, this may represent an additional potential risk for kidney integrity, which is already compromised by the cholestatic process itself (cholemic nephropathy). Therefore, in the present work, we studied the renal damage caused by ANIT-induced cholestasis and whether it is aggravated or, on the contrary, counteracted by HGF; if the latter holds, the involvement of its antioxidant properties will be ascertained. ANIT-induced cholestatic deleterious renal effects were corroborated by the presence of urine bile salts, impairment of renal function, and the alterations of renal damage markers, such as HSP72, creatinine clearance, and albuminuria. HGF fully reverted all these, and the cast formation in the tubules was significantly decreased. These findings were associated with the control of renal oxidative stress. In summary, despite HGF enhancing the overload of potentially harmful biliary constituents that the kidney should remove from the bloodstream as an alternative depuration organ in cholestasis, it simultaneously protects the kidney from this damage by counteracting the prooxidant effects resulting from this harmful exposure.

Conditioned place preference for cocaine and methylphenidate in female mice from lines selectively bred for high voluntary wheel-running behavior.

Behavioral addictions can come in many forms, including overeating, gambling and overexercising. All addictions share a common mechanism involving activation of the natural reward circuit and reinforcement learning, but the extent to which motivation for natural and drug rewards share similar neurogenetic mechanisms remains unknown. A unique mouse genetic model in which four replicate lines of female mice were selectively bred (>76 generations) for high voluntary wheel running (High Runner or HR lines) alongside four non-selected control (C) lines were used to test the hypothesis that high motivation for exercise is associated with greater reward for cocaine (20 mg/kg) and methylphenidate (10 mg/kg) using the conditioned place preference (CPP) test. HR mice run ~three times as many revolutions/day as C mice, but the extent to which they have increased motivation for other rewards is unknown. Both HR and C mice displayed significant CPP for cocaine and methylphenidate, but with no statistical difference between linetypes for either drug. Taken together, results suggest that selective breeding for increased voluntary running has modified the reward circuit in the brain in a way that increases motivation for running without affecting cocaine or methylphenidate reward.

HGF induces protective effects in α-naphthylisothiocyanate-induced intrahepatic cholestasis by counteracting oxidative stress.

Cholestasis is a clinical syndrome common to a large number of hepatopathies, in which either bile production or its transit through the biliary tract is impaired due to functional or obstructive causes; the consequent intracellular retention of toxic biliary constituents generates parenchyma damage, largely via oxidative stress-mediated mechanisms. Hepatocyte growth factor (HGF) and its receptor c-Met represent one of the main systems for liver repair damage and defense against hepatotoxic factors, leading to an antioxidant and repair response. In this study, we evaluated the capability of HGF to counteract the damage caused by the model cholestatic agent, α-naphthyl isothiocyanate (ANIT). HGF had clear anti-cholestatic effects, as apparent from the improvement in both bile flow and liver function test. Histology examination revealed a significant reduction of injured areas. HGF also preserved the tight-junctional structure. These anticholestatic effects were associated with the induction of basolateral efflux ABC transporters, which facilitates extrusion of toxic biliary compounds and its further alternative depuration via urine. The biliary epithelium seems to have been also preserved, as suggested by normalization in serum GGT levels, CFTR expression and cholangyocyte primary cilium structure our results clearly show for the first time that HGF protects the liver from a cholestatic injury.

Resource Utilization and Costs Associated With Percutaneous Coronary Intervention When Treating Patients With Acute Coronary Syndrome in the Mexican National Institute of Cardiology "Ignacio Chávez".

BACKGROUND: The treatment of acute coronary syndrome (ACS) using percutaneous coronary intervention (PCI) is a frequent intervention with a high economic impact. OBJECTIVE: This study investigates the resource use and cost of PCI in Mexico where heart disease is a leading cause of death, and a large segment of the population does not have formal healthcare coverage. METHODS: This retrospective observational study obtained resource utilization data from patient files and itemized costs from the pharmacy registry at the National Institute of Cardiology. Patients were aged >18 years, diagnosed with ACS, and treated with PCI and secondary prophylaxis with aspirin plus clopidogrel or prasugrel. Patients had a follow-up of >12 months at the institute. Statistical analysis was descriptive. RESULTS: The sample included 156 patients (mean age: 58.66 years; male: 77.9%). Patients were diagnosed with ST segment elevation myocardial infarction (STEMI), non-ST segment elevation myocardial infarction, and unstable angina 64.9%, 27.2%, and 7.9%, respectively. The mean (standard deviation [SD]) total medical cost was estimated to be $145 677 ($98 326) Mexican pesos 2018. The highest category of spending was surgical materials (mean [SD]: $47 834 [$32 569], comprising 32.8% of total costs); medications and access to the operating room represented 14.2% and 11.8%, respectively. Mean (SD) hospital stay was 9.07 (6.2) days; for the 11.5% of patients admitted to the intensive care unit, the mean (SD) stay was 4.61 (2.06) days. The mean cost of standard hospitalization was $12 572, or 8.6% of spending; intensive care unit hospitalization comprised 17.7% of total costs (mean: $25 802). The cost of the intervention is subsidized up to 95% for patients with a low social economic status, with the exception of surgical materials such as stents. This results in the highest burden component of the intervention being placed on the patient and not the institute. CONCLUSION: The mean cost per patient shows that PCI is an expensive procedure in Mexico. A lack of subsidies for surgical equipment places a high economic burden on the patient and represents a barrier of access for a vulnerable population that likely increases mortality and morbidity rates in those patients unable to pay for treatment and a potential high burden of debt for those who do pay.

Cholangiocyte death in ductopenic cholestatic cholangiopathies: Mechanistic basis and emerging therapeutic strategies.

Among hepatic diseases, cholestatic ductopenic cholangiopathies are poorly studied, and they are rarely given the importance they deserve, especially considering their high incidence in clinical practice. Although cholestatic ductopenic cholangiopathies have different etiologies and pathogenesis, all have the same target (the cholangiocyte) and similar mechanistic basis of cell death. Cholestatic cholangiopathies are characterized, predominantly, by obstructive or functional damage in the biliary epithelium, resulting in an imbalance between proliferation and cholangiocellular death; this leads to the progressive disappearance of bile ducts, as has been shown to occur in primary sclerosing cholangitis, primary biliary cholangitis, low-phospholipid-associated cholelithiasis syndrome, cystic fibrosis-related liver disease, and drug-induced ductopenia, among other biliary disorders. This review summarizes the features of the more common ductopenic syndromes and the cellular mechanisms involved in cholengiocellular death, with focus on the main forms of cholangiocyte death described so far, namely apoptosis, autophagy, necrosis, and necroptosis. It also emphasizes the importance to study in depth the molecular mechanisms of cholengiocyte death to make possible to counteract them with therapeutic purposes. These therapeutic strategies are limited in number and efficacy at present, and this is why it is important to find complementary, safe strategies to stimulate cholangiocellular proliferation in order favor bile duct replenishment as well. Successful in finding appropriate treatments would prevent the patient from having liver transplantation as the only therapeutic alternative.

Reduced non-bicarbonate skeletal muscle buffering capacity in mice with the mini-muscle phenotype.

Muscle pH decreases during exercise, which may impair function. Endurance training typically reduces muscle buffering capacity as a result of changes in fiber-type composition, but existing comparisons of species that vary in activity level are ambiguous. We hypothesized that high-runner (HR) lines of mice from an experiment that breeds mice for voluntary wheel running would have altered muscle buffering capacity as compared with their non-selected control counterparts. We also expected that 6 days of wheel access, as used in the selection protocol, would reduce buffering capacity, especially for HR mice. Finally, we expected a subset of HR mice with the 'mini-muscle' phenotype to have relatively low buffering capacity as a result of fewer type IIb fibers. We tested non-bicarbonate buffering capacity of thigh muscles. Only HR mice expressing the mini-muscle phenotype had significantly reduced buffering capacity, females had lower buffering capacity than males, and wheel access had no significant effect.