Angelman Syndrome Protein Ube3a Regulates Synaptic Growth and Endocytosis by Inhibiting BMP Signaling in Drosophila

Altered expression of the E3 ubiquitin ligase UBE3A, which is involved in protein degradation through the proteasome-mediated pathway, is associated with neurodevelopmental and behavioral defects observed in Angelman syndrome (AS) and autism. However, little is known about the neuronal function of UBE3A and the pathogenesis of UBE3A-associated disorders. To understand the in vivo function of UBE3A in the nervous system, we generated multiple mutations of ube3a, the Drosophila ortholog of UBE3A. We found a significantly increased number of total boutons and satellite boutons in conjunction with compromised endocytosis in the neuromuscular junctions (NMJs) of ube3a mutants compared to the wild type. Genetic and biochemical analysis showed upregulation of bone morphogenetic protein (BMP) signaling in the nervous system of ube3a mutants. An immunochemical study revealed a specific increase in the protein level of Thickveins (Tkv), a type I BMP receptor, but not other BMP receptors Wishful thinking (Wit) and Saxophone (Sax), in ube3a mutants. Ube3a was associated with and specifically ubiquitinated lysine 227 within the cytoplasmic tail of Tkv, and promoted its proteasomal degradation in Schneider 2 cells. Negative regulation of Tkv by Ube3a was conserved in mammalian cells. These results reveal a critical role for Ube3a in regulating NMJ synapse development by repressing BMP signaling. This study sheds new light onto the neuronal functions of UBE3A and provides novel perspectives for understanding the pathogenesis of UBE3A-associated disorders.     

Out of Pocket Expenditure for Hospitalization among Below Poverty Line Households in District Solan,Himachal Pradesh,India, 2013

Introduction

Health insurance schemes, like Rashtriya Swasthya Bima Yojana (RSBY), should provide financial protection against catastrophic health costs by reducing out of pocket expenditure (OOPE) for hospitalizations. We estimated and compared the proportion and extent of OOPE among below poverty line (BPL) families beneficiaries and not beneficiaries by RSBY during hospitalizations in district Solan, H.P., India, 2013.

Methods

We conducted a cross sectional survey among hospitalized BPL families in the beneficiaries and non-beneficiaries groups. We compared proportion incurring OOPE and its extent during hospitalization, pre/post-hospitalization periods in different domains.

Results

Overall, proportion of non-beneficiaries who incurred OOPE was higher than the beneficiaries but it was not statistically significant (87.2% vs. 80.9%). The median overall OOPE was $39 (Rs 2567) in the non-beneficiaries group as compared to $ 11 (Rs 713) in the beneficiaries group (p<0.01). Median expenditure on in house and out house drugs and consumables was $ 23 (Rs 1500) in the non beneficiaries group as compared to nil in the beneficiaries group (p<0.01). Non-beneficiary status was significantly associated [OR: 2.4 (1.3–4.3)] with OOPE above median independently and also after adjusting for various covariates.

Conclusion

RSBY has decreased the extent of OOPE among the beneficiaries; however OOPE was incurred mainly due to purchase of drugs from outside the health facility. The treatment seeking behaviour in beneficiaries group has improved among comparatively older group with chronic conditions. RSBY has enabled beneficiaries to get more facilities such as drugs, consumables and diagnostics from the health facility.     

Why individual thermo sensation and pain perception varies? Clue of disruptive mutations in TRPVs from 2504 human genome data

Every individual varies in character and so do their sensory functions and perceptions. The molecular mechanism and the molecular candidates involved in these processes are assumed to be similar if not same. So far several molecular factors have been identified which are fairly conserved across the phylogenetic tree and are involved in these complex sensory functions. Among all, members belonging to Transient Receptor Potential (TRP) channels have been widely characterized for their involvement in thermo-sensation. These include TRPV1 to TRPV4 channels which reveal complex thermo-gating behavior in response to changes in temperature. The molecular evolution of these channels is highly correlative with the thermal response of different species. However, recent 2504 human genome data suggest that these thermo-sensitive TRPV channels are highly variable and carry possible deleterious mutations in human population. These unexpected findings may explain the individual differences in terms of complex sensory functions.     

Sirtuin 1 and 7 mediate resveratrol-induced recovery from hyper-anxiety in high-fructose-fed prediabetic rats

Hyperglycaemia in diabetes is either caused by reduced availability of insulin (type 1 diabetes, T1D) or insulin resistance to the cells (type 2 diabetes, T2D). In recent years, the prevalence of T2D has increased to an alarming proportion, encompassing 95% of the total diabetic burden, probably due to economy-driven changes in lifestyle. Recent epidemiological studies show comorbid depression, anxiety and related mental illness. To explore the molecular mechanisms underlying this comorbid conditions, we used Sprague–Dawley rats on high-fructose diet for 8 weeks to induce prediabetic condition. Rats with this metabolic syndrome also showed hyper-anxiety when they were subjected to anxiety-related behavioural assays. Rats were administered with resveratrol, an activator of sirtuins, and metformin, a standard antidiabetic drug, simultaneously with fructose. We observed that resveratrol was more effective in protecting from both the metabolic (prediabetic) and affective (anxiety) disorders than metformin. Molecular studies showed that recovery was associated with the upregulation of few nuclear sirtuins that act epigenetically – Sirt 1 and 7, which were significantly attenuated in the striatum of prediabetic rats. In conclusion, our study showed that hyper-anxiety associated with prediabetic condition is ameliorated by resveratrol through modulation of sirtuins, which is more or less similar to metformin.     

<Emphasis Type="Italic">In Vitro</Emphasis> Anti-inflammatory and Antimicrobial Activities of Azithromycin After Loaded in Chitosan- and Tween 20-Based Oil-in-Water Macroemulsion for Acne Management

The objectives of the current investigation are (1) to prepare and characterize (particle size, surface charge (potential zeta), surface morphology by transmission electron microscopy, drug content, and drug release) the azithromycin (AZM, 100 mg)-loaded oil-in-water (o/w) macroemulsion, (2) to assess the toxicity of macroemulsion with or without AZM using RBC lysis test in comparison with AZM in phosphate buffer solution of pH 7.4, (3) to compare the in vitro antimicrobial activity (in Escherichia coli using zone inhibition assay) of AZM-loaded macroemulsion with its aqueous solution, and (4) to assess the in vitro anti-inflammatory effect (using egg albumin denaturation bioassay) of the AZM-loaded macroemulsion in comparison with diclofenac sodium in phosphate buffer solution of pH 7.4. The AZM-loaded macroemulsion possessed the dispersed oil droplets with a mean diameter value of 52.40?±?1.55 μm. A reversal in the zeta potential value from negative (?2.16?±?0.75 mV) to positive (+6.52?±?0.96 mV) was noticed when AZM was added into the macroemulsion. At a 1:5 dilution ratio, 2.06?±?0.03 mg of drug was released from macroemulsion followed by 1.01?±?0.01 and 0.25?±?0.08 mg, respectively, for 1:10 and 1:40 dilution ratios. Antimicrobial activity maintenance and significant reduction of RBC lysis property were noticed for AZM after loaded in the macroemulsion. However, an increment in the absorbance values for emulsion-treated samples in comparison to the control samples was noticed in the anti-inflammatory test. This speculates the potential of the AZM-loaded emulsion to manage inflammatory conditions produced at Acne vulgaris.     

Expression of Putative Stem Cell Marker,Hepatocyte Nuclear Factor 4 Alpha,in Mammary Gland of Water Buffalo

Buffaloes account for more than 56% of total milk production in India. Cyclic remodeling of mammary glands of human, mice, cow, sheep, and goat is determined by mammary stem cells. It is logical to assume that buffalo mammary gland will have mammary stem/progenitor cells. Thus far, no report exists on identification of buffalo mammary stem cells. Hepatocyte nuclear factor 4 alpha (HNF4A) is a candidate marker for hepatic progenitor cells and has recently been suggested as a marker of bovine mammary stem/progenitor cells. We hypothesized that (1 Pasha TN, Hayat Z. Present situation and future perspective of buffalo production in Asia. J Anim Plant Sci 2012; 22(3 supple.):250–256. [Google Scholar]) HNF4A identifies putative buffalo mammary stem/progenitor cells and (2 NDDB. National Dairy Development Board. 2015. http://www.nddb.org/English/Statistics/Pages/Milk-Production.aspx. Accessed May 10, 2015. [Google Scholar]) the number of HNF4A-positive cells increases during mastitis. Sixteen buffalo mammary samples were collected from a local slaughterhouse. Hematoxylin and eosin staining were performed on 5-micron thick sections and on the basis of gross examination and histomorphology of the mammary glands, physiological stages of the animals were estimated as non-lactating (n = 4), mastitis (n = 9), and prepubertal (n = 3). In total, 24048 cells were counted (5–10 microscopic fields/animal; n = 16 animals) of which, 40% cells were mammary epithelial cells (MEC) and 60% cells were the stromal cells. The percentage of MEC in non-lactating animals was higher compared to mastitic animals (47.3% vs. 37.3%), which was likely due to loss of MEC in mastitis. HNF4A staining was observed in nuclei of MEC of ducts, alveoli, and stromal cells. Basal location and low frequency of HNF4A-positive MEC (ranges from 0.4–4.5%) were consistent with stem cell characteristics. Preliminary study showed coexpression of HNF4A with MSI1 (a mammary stem cell marker in sheep), suggesting HNF4A was likely to be a putative mammary stem/progenitor cell marker in buffalo. HNF4A-positive MEC (basal and luminal; light and dark stained) tended to be higher in non-lactating than the mastitic animals (8.73 ± 1.71% vs. 4.29 ± 1.19%; P = 0.07). The first hypothesis that HNF4A identify putative mammary stem/progenitor cells was confirmed but the second hypothesis that the number of mammary stem/progenitor cells decreases during mastitis was unsupported. This is the first report outlining the expression of HNF4A and identification of putative mammary stem/progenitor cells in buffalo mammary gland.     

Molecular Characterization of Pro-Inflammatory Cytokines Interleukin-1β and Interleukin-8 in Asian Elephant (Elephas maximus)

Interleukin (IL)-1β and IL-8 are pro-inflammatory cytokines produced primarily by monocytes and macrophages in response to a variety of microbial and nonmicrobial agents. As yet, no molecular data have been reported for IL-1β and IL-8 of the Asian elephant. In the present study, we have cloned and sequenced the cDNA encoding IL-1β and IL-8 of the Asian elephant. The open reading frame (ORF) of Asian elephant IL-1β is 789 bp in length, encoded a propeptide of 263 amino acid polypeptide. The predicted protein revealed the presence of IL-1 family signature motif and an ICE cut site. Whereas, IL-8 contained 321 bp of open reading frame. Interestingly, the predicted protein sequence of 106 aa, contains an ELR motif immediately upstream of the CQC residues, common in all vertebrate IL-8 molecules. Identity levels of the nucleic acid and deduced amino acid sequences of Asian elephant IL-1β ranged from 68.48 (Squirrel monkey) to 98.57% (African elephant), and 57.78 (Sheep) to 98.47% (African elephant), respectively, whereas that of IL-8 ranged from 72.9% (Human) to 87.8% (African elephant), and 63.2 (human, gorilla, chimpanzee) to 74.5% (African elephant, buffalo), respectively. The phylogenetic analysis based on deduced amino acid sequenced showed that the Asian elephant IL-1β and IL-8 were most closely related to African elephant. Molecular characterization of these two cytokines, IL-1β and IL-8, in Asian elephant provides fundamental information necessary to progress the study of functional immune responses in this animal and gives the potential to use them to manipulate the immune response as recombinant proteins.