Synthesis,biological activities,and pharmacokinetics studies of a mutual prodrug of aceclofenac and paracetamol

An ester-based mutual prodrug (aceclofenac–paracetamol; AC-PR) was synthesized (one-pot method) with an aim of improving the therapeutic index through prevention of gastrointestinal irritation and bleeding that is associated with aceclofenac. The release of aceclofenac and paracetamol from the ester prodrug (AC-PR) was studied by reverse phase HPLC in hydrochloric acid buffer (pH 1.2), phosphate buffer (pH 7.4), 80 % v/v human plasma, 10 % w/v rat intestinal homogenate and 10 % w/v rat liver homogenate (pH 7.4). The prodrug showed negligible hydrolysis at pH 1.2 as compared to pH 7.4, suggesting that very less of the prodrug would hydrolyze in stomach, but would release the parent drugs at pH 7.4 in adequate amounts. The prodrug showed enhanced anti-inflammatory activity and significant protection against acetic acid-induced writhings (analgesic activity) as compared to that of aceclofenac. Further, the prodrug produced reduced number of ulcers as compared to that of the parent drug. These results suggest that the synthesized mutual prodrug (AC-PR) is better in terms of activity and GIT toxicity than the parent drug.     

Pre-pubertal exposure of cytarabine-induced testicular atrophy,impaired spermatogenesis and germ cell DNA damage in SD rats

Context: Cytarabine (Ara-C) is an effective chemotherapeutic drug for the treatment of acute leukaemias. It inhibits the DNA synthesis and repair, thereby causes cytotoxicity in the proliferating cells.

Objective: This study was aimed to investigate the effects of pre-pubertal exposure of Ara-C on testesticular development in juvenile SD rats and their function at puberty.

Materials and methods: Ara-C was injected at the doses of 50, 100 and 200?mg/kg/day from postnatal day (PND) 29–42 (14 days) by intraperitoneal (i.p.) route. Half of the animals were sacrificed on PND 43 and remaining on PND 70. End points of the evaluation included gross pathological examination, histomorphometric analysis, sperm count and sperm head morphology, cell proliferation and DNA damage as well as apoptosis analysis.

Results: Ara-C treatment significantly decreased food and water intake, weight gain, testes and epididymis weight and increased histological alterations in the seminiferous tubule. Furthermore, Ara-C treatment significantly decreased the PCNA-positive cells and sperm count in a dose-dependent manner. Ara-C treatment also increased the DNA damage and apoptosis in testes and sperm as evident from the comet and TUNEL assays results.

Discussion: The present study results clearly indicated that Ara-C treatment impaired spermatogenesis and adversely affects the testicular development and its function in rats by reducing the germ cell proliferation and the inducing DNA damage and apoptosis.     

Phosphoproteomic Analysis Reveals Regulatory Mechanisms at the Kidney Filtration Barrier

Diseases of the kidney filtration barrier are a leading cause of ESRD. Most disorders affect the podocytes, polarized cells with a limited capacity for self-renewal that require tightly controlled signaling to maintain their integrity, viability, and function. Here, we provide an atlas of in vivo phosphorylated, glomerulus-expressed proteins, including podocyte-specific gene products, identified in an unbiased tandem mass spectrometry–based approach. We discovered 2449 phosphorylated proteins corresponding to 4079 identified high-confidence phosphorylated residues and performed a systematic bioinformatics analysis of this dataset. We discovered 146 phosphorylation sites on proteins abundantly expressed in podocytes. The prohibitin homology domain of the slit diaphragm protein podocin contained one such site, threonine 234 (T234), located within a phosphorylation motif that is mutated in human genetic forms of proteinuria. The T234 site resides at the interface of podocin dimers. Free energy calculation through molecular dynamic simulations revealed a role for T234 in regulating podocin dimerization. We show that phosphorylation critically regulates formation of high molecular weight complexes and that this may represent a general principle for the assembly of proteins containing prohibitin homology domains.The kidney filter consists of three layers: fenestrated endothelial cells, the glomerular basement membrane, and podocytes.¹ Damage to any of these compartments becomes clinically evident as proteinuria and the development of kidney disease.² Of particular importance for the regulation of podocyte biology through signaling is the slit diaphragm, a specialized intercellular junction that bridges the 40-nm gap in between foot processes of neighboring podocytes. It also serves as a signaling platform regulating podocyte function. Mutations in genes encoding for components of the slit diaphragm, such as nephrin,³ podocin,⁴ CD2AP,⁵ and TRPC6,^6,7 are important causes of genetic forms of proteinuria. Alteration of these proteins results in defective signaling causing podocyte dysfunction, progressive glomerulosclerosis, and kidney failure. The slit diaphragm protein complex is a lipid-multiprotein supercomplex.⁸ Of central importance to the integrity and function of the protein complex is the prohibitin homology (PHB) domain protein podocin,⁹ which forms multimeric complexes and is required to control signal transduction through associated transmembrane proteins.^10,11Signaling processes governing podocyte function, integrity, and survival largely depend on signaling processes involving phosphorylation.^12,13 Comprehensive analyses of the signaling events in podocytes in vivo have been hampered by the fact that interference with these signaling cascades by genetic deletion often results in massively disrupted and dysfunctional podocytes. One of the primary aims of this study was to use phosphoproteomics to analyze thousands of phosphorylation sites in native murine glomeruli within single samples. Within this study, we show that this approach allows the introduction of new concepts into signaling processes at the kidney filtration barrier.     

Evaluation of aqueous extract of Murraya koenigii in unilateral renal ischemia reperfusion injury in rats

Aim:

The aqueous extract of leaves of Murraya koenigii was studied for its renoprotective potential against unilateral renal ischemia reperfusion (RIR) injury in male Wistar rats.

Materials and Methods:

Healthy adult male Wistar rats were divided into five groups (n = 8) and were treated with 200 mg/kg., p.o. of aqueous extract of M. koenigii (AEMK) for 30 days to assess both preventive and curative effects of AEMK. Except Group I, RIR was induced to all the groups by clamping the left renal artery using artery clamp for 1 h followed by reperfusion by removing the clamp. Groups II and III underwent RIR at 30^th day whereas RIR was induced in Groups IV and V at 1^st day of treatment schedule. Biochemical parameters (serum creatinine, blood urea nitrogen, serum total protein and serum Na⁺), urinary parameters (urine output, urinary creatinine, urinary urea, urinary total protein, urinary Na⁺), in vivo anti-oxidants, renal myeloperoxidase (MPO) activity and histopathology of kidneys were monitored. Statistical significance was set at P < 0.05.

Results:

Rats were treated with AEMK significantly (P < 0.05) restored the serum and urinary parameters with significant (P < 0.05) improvement in endogenous anti-oxidants such as superoxide dismutase, catalase and reduced glutathione and decreased levels of malondialdehyde and renal MPO when compared with the control groups. Histopathological examination also supported the biochemical and urinary tests.

Conclusions:

Aqueous extract of M. koenigii possesses both preventive and curative effects against RIR injury.KEY WORDS: Aqueous extract, ischemia reperfusion injury, kidney, Murraya koenigii     

Malnutrition after pancreatic enzyme replacement therapy in chronic pancreatitis: Risk factors in real world practice

BackgroundRCTs that have shown improvement in coefficient of fat absorption with pancreatic enzyme replacement therapy (PERT) have seldom evaluated the impact on overall nutritional status.ObjectiveIn this study we evaluated factors responsible for persistence of malnutrition after PERT.MethodsIn this cross-sectional observational study, patients were enrolled based on predefined enrolment criteria. Patients were divided into those taking PERT regularly (Group A), irregularly (Group B) and not taking (Group C) for at least 3 months. Comprehensive evaluation of anthropometric measurements, nutritional assessment and dietary intake was performed. Malnutrition was measured using the Subjective Global Assessment (SGA) tool. Relationship between PERT status, dietary intake and nutritional status were evaluated using standard statistical methods. Logistic regression was performed to identify factors associated with persistence of malnutrition after PERT.Results377 patients with CP and 50 controls were included. 95 (25.2%) patients with CP were in Group A, 106 (28.1%) in Group B and 176 (46.7%) in Group C. 130 (34.5%) patients were malnourished, of which 76 (58.5%) were continuing PERT. There were no differences in clinical and biochemical nutritional markers between Groups A, B, and C. Calorie deficit and daily intake of calorie, protein, carbohydrates and fats were not different between those with and without PERT, but was significantly less in those with malnutrition. Logistic regression demonstrated inadequate dietary intake as independent risk factor for persistence of malnutrition.ConclusionEven though PERT is effective in PEI, comprehensive nutritional assessment, personalized nutritional counselling and therapy along with PERT is mandatory.     

Challenges in optimizing sample preparation and LC‐MS/MS conditions for the analysis of carglumic acid,an N‐acetyl glutamate derivative in human plasma

This paper describes a systematic approach to overcoming challenges in developing a robust and selective liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) method for reliable and precise determination of carglumic acid in human plasma. Sample extraction was tested on several reversed‐phase solid‐phase extraction (SPE) sorbents with different chemistries, such as hydrophobic C18, hydrophilic‐lipophilic balance, and mixed‐mode cation and anion exchange. The best recovery under the optimized extraction conditions was obtained with Oasis MAX (30 mg, 1cc) mixed‐mode anion exchange (~ 50%) cartridge, compared to other sorbents from 100 μL plasma sample. Complete analytical separation of carglumic acid and carglumic acid‐13C5 15N as an internal standard (IS) from endogenous plasma components was achieved on ACE 5CN (150 × 4.6 mm, 5 µm) column under isocratic conditions using acetonitrile:methanol (50:50, v/v) ? 0.1% acetic acid in water [80:20, v/v] as the mobile phase. The deprotonated precursor → product ion transitions for carglumic acid (189/146) and IS (195/152) were monitored in the negative ionization mode on a triple quadrupole mass spectrometer. The regression curves were linear over a concentration range of 6.00‐6000 ng/mL (r² ≥ 0.9987). Matrix effect was evaluated in terms of IS‐normalized matrix factors, which ranged from 0.95 to 1.01 across four quality control levels. Intra‐ and inter‐batch accuracy and precision, and the stability of carglumic acid in spiked plasma samples were assessed under different conditions. The method was applied to assess the pharmacokinetics of 100 mg/kg body weight carglumic acid in a healthy Indian subject. Copyright © 2015 John Wiley & Sons, Ltd.     

Chronic low-level arsenic exposure reduces lung function in male population without skin lesions

Objectives

The respiratory effects of chronic low-level arsenic exposure from groundwater have been investigated in West Bengal, India.

Methods

The participants (834 non-smoking adult males) were subdivided in two groups: an arsenic-exposed group (n = 446, mean age 35.3 years) drinking arsenic-contaminated groundwater (11–50 μg/L) and a control group of 388 age-matched men drinking water containing <10 μg/L of arsenic. Arsenic in water samples was measured by atomic absorption spectroscopy. The prevalence of respiratory symptoms was documented by structured, validated questionnaire. Pulmonary function test (PFT) was assessed by portable spirometer.

Results

Compared with control, the arsenic-exposed subjects had higher prevalence of upper and lower respiratory symptoms, dyspnea, asthma, eye irritation and headache. Besides, 20.6 % of arsenic-exposed subjects had lung function deficits (predominantly restrictive and combined types) compared with 13.6 % of control (p < 0.05). A positive association was observed between arsenic concentration in drinking water and the prevalence of respiratory symptoms, while a negative association existed between arsenic level and spirometric parameters.

Conclusions

The findings suggest that even low-level arsenic exposure has deleterious respiratory effects.     

Integrity of medial temporal structures may predict better improvement of spatial neglect with prism adaptation treatment

Prism adaptation treatment (PAT) is a promising rehabilitative method for functional recovery in persons with spatial neglect. Previous research suggests that PAT improves motor-intentional “aiming” deficits that frequently occur with frontal lesions. To test whether presence of frontal lesions predicted better improvement of spatial neglect after PAT, the current study evaluated neglect-specific improvement in functional activities (assessment with the Catherine Bergego Scale) over time in 21 right-brain-damaged stroke survivors with left-sided spatial neglect. The results demonstrated that neglect patients’ functional activities improved after two weeks of PAT and continued improving for four weeks. Such functional improvement did not occur equally in all of the participants: Neglect patients with lesions involving the frontal cortex (n?=?13) experienced significantly better functional improvement than did those without frontal lesions (n?=?8). More importantly, voxel-based lesion-behavior mapping (VLBM) revealed that in comparison to the group of patients without frontal lesions, the frontal-lesioned neglect patients had intact regions in the medial temporal areas, the superior temporal areas, and the inferior longitudinal fasciculus. The medial cortical and subcortical areas in the temporal lobe were especially distinguished in the “frontal lesion” group. The findings suggest that the integrity of medial temporal structures may play an important role in supporting functional improvement after PAT.     

Society of Abdominal Radiology (SAR) and European Society of Urogenital Radiology (ESUR) joint consensus statement for MR imaging of placenta accreta spectrum disorders

This study was conducted in order to establish the joint Society of Abdominal Radiology (SAR) and European Society of Urogenital Radiology (ESUR) guidelines on placenta accreta spectrum (PAS) disorders and propose strategies to standardize image acquisition, interpretation, and reporting for this condition with MRI. The published evidence-based data and the opinion of experts were combined using the RAND–UCLA Appropriateness Method and formed the basis for these consensus guidelines. The responses of the experts to questions regarding the details of patient preparation, MRI protocol, image interpretation, and reporting were collected, analyzed, and classified as “recommended” versus “not recommended” (if at least 80% consensus among experts) or uncertain (if less than 80% consensus among experts). Consensus regarding image acquisition, interpretation, and reporting was determined using the RAND–UCLA Appropriateness Method. The use of a tailored MRI protocol and standardized report was recommended. A standardized imaging protocol and reporting system ensures recognition of the salient features of PAS disorders. These consensus recommendations should be used as a guide for the evaluation of PAS disorders with MRI. • MRI is a powerful adjunct to ultrasound and provides valuable information on the topography and depth of placental invasion. • Consensus statement proposed a common lexicon to allow for uniformity in MRI acquisition, interpretation, and reporting of PAS disorders. • Seven MRI features, namely intraplacental dark T2 bands, uterine/placental bulge, loss of low T2 retroplacental line, myometrial thinning/disruption, bladder wall interruption, focal exophytic placental mass, and abnormal vasculature of the placental bed, reached consensus and are categorized as “recommended” for diagnosing PAS disorders.     

CCR9 signaling in dendritic cells drives the differentiation of Foxp3+ Tregs and suppresses the allergic IgE response in the gut

The chemokine receptor CCR9 and its only known ligand CCL25 play an important role in gut inflammation and autoimmune colitis. The function of CCR9-CCL25 in the migration of immune cells is well characterized. However, its role in the immune cell differentiation is mostly not known. Using dextran sodium sulfate (DSS)-induced gut inflammation model, we showed that CCR9⁺ dendritic cells (DCs) specifically CD11b⁻CD103⁺ DCs were significantly increased in the gut-associated lymphoid tissues (GALT) compared to control mice. These CCR9⁺ DCs express lower MHC II and CD86 molecules and had regulatory surface markers (FasL and latency-associated peptide, LAP) in the GALT. In the presence of CCL25, CCR9⁺ DCs promoted in vitro differentiation of Foxp3⁺ regulatory CD4⁺ T cells (Tregs). CCL25-induced differentiation of Tregs was due to intrinsic signaling in the DCs but not through CD4⁺ T cells, which was driven by the production of thymic stromal lymphopoietin (TSLP) and not IL-10. Furthermore, adoptive transfer of CCR9⁺ DCs in C57BL/6 mice promoted Tregs but reduced the Th17 cells in the GALT, and also suppressed the OVA-specific gut-allergic response. Our results suggest CCR9⁺ DCs have a regulatory function and may provide a new cellular therapeutic strategy to control gut inflammation and allergic immune reaction.