Distinct bactericidal activities of bovine lactoferrin peptides LFampin 268-284 and LFampin 265-284: Asp-Leu-Ile makes a difference.

Two lactoferrampin (LFampin) peptides derived from bovine lactoferrin were compared with respect to their bactericidal activities. LFampin 265-284 killed a set of Gram-positive bacteria that were resistant to LFampin 268-284. The presence of 265Asp-Leu-267Ile did not simply lead to an overall increased potency, since higher concentrations of LFampin 265-284 than LFampin 268-284 were needed to kill the Gram-negative bacteria that were tested. The Asp-Leu-Ile sequence enhances the propensity of LFampin to adopt an alpha-helix, as shown by circular dichroism spectroscopy. These results suggest that the helical conformation of the peptide is an important determinant of the susceptibility of Gram-positive bacteria.     

The Effects of Synbiotic Supplementation on Glucose Metabolism and Lipid Profiles in Patients with Diabetes: a Systematic Review and Meta-Analysis of Randomized Controlled Trials

Probiotics and Antimicrobial Proteins - Although several studies have evaluated the effect of synbiotic intake on metabolic profiles in patients with diabetes, findings are inconsistent. This...     

Effect of low‐level laser therapy and oxytocin on osteoporotic bone marrow‐derived mesenchymal stem cells

Postmenopausal osteoporosis (OP) is a major concern for public health. Low‐level laser therapy (LLLT) has a positive effect on the health of bone marrow mesenchymal stem cells (BMMSCs). The purpose of this study is to evaluate the influence of LLLT and oxytocin (OT) incubation—individually and in combination—on osteoporotic BMMSCs in ovariectomized rats. Twelve female rats were randomized into two groups to undergo either a sham surgery (sham group) or ovariectomy‐induced osteoporosis (OVX group). MSCs harvested from the BM of healthy and OVX rats underwent culture expansion. There were five groups. In Groups one (sham‐BMMSC) and two (OVX‐BMMSC) the cells were held in osteogenic condition medium without any intervention. In the group three (OT), OT incubation with optimum dose was performed for 48 h (two times, 10^?12 molar). In Group four, laser‐treated‐OVX‐BMMSCs were treated with optimum protocol of LLLT (one time, 1.2 J/cm²). In Group five (laser + OT group), the OT incubation plus the laser irradiation was performed. The biostimulatory effect of LLLT is demonstrated by a significant increase in the viability of OVX‐BMMSCs, cell cycle, and extracellular levels of Transforming growth factor beta (TGF‐β), insulin‐like growth factor‐I (IGF‐I), and Alkaline phosphatase (ALP) compared to control OVX‐BMMSCs and/or the sham group. OT incubation and laser + OT incubation have a positive effect on OVX‐BMMSCs. However, LLLT is more effective statistically. We conclude that LLLT significantly improved cell viability, enhanced the osteogenic potential of the OVX‐BMMSCs, and increased the extracellular levels of the TGF‐β, IGF‐I, and ALP.     

Incidence of metallo-beta-lactamase-producing <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis> isolates from hospital setting in Pakistan

The aim of this study was monitoring and surveillance in different wards of the PIMS hospital, Islamabad, to understand emerging challenges of antibiotic resistance in particular association with most virulent serotypes of Klebsiella pneumoniae. The study was conducted during March 2015 to September 2015. The study showed that rate of isolation of K. pneumoniae was 37% (103 positives out of a total of 277 clinical samples) and 7.7% (8) were phenotypically and genotypically confirmed to be metallo-β-lactamase resistant (carbapenem resistant) and all of them were multidrug resistant (MDR). These carbapenem-resistant isolates were isolated from blood, endotracheal tubes, and pus. Molecular screening for the presence of integrons indicated that distribution of class I integrons (87.5% of carbapenem-resistant K. pneumoniae isolates) was higher than class II integrons (1.25%) among given isolates. The study indicated that exposure of metallo-beta-lactamase-producing strains through hospitalizations increases the chances of spread of MDR pathogens. There is an urgent need for effective surveillance and monitoring strategies to control the spread of extremely resistant K. pneumoniae implicated in nosocomial infections leading to the increased health burden and enforcement of policy guideline on appropriate antibiotics usage.     

First genetic evidence for the presence of the rumen fluke Paramphistomum epiclitum in Pakistan

More than 70 species of the Superfamily Paramphistomoidea, have been identified in ruminants in different parts of the world. Most are pathogenic, causing amphistomosis. Adult flukes within this family have a predilection for the forestomach (rumen) or bile duct of the liver, where they may cause epithelial damage. Identification of adult Paramphistomum, Calicophoron, Gastrothylax and Fischoederius at the species level based on morphology requires specialised expertise, whereas molecular genetic marker analysis is more precise and transferable. In the present study, we performed molecular characterisation of twenty seven adult flukes collected from the forestomachs of buffalo, cattle and goats in the Punjab province of Pakistan. PCR and sequencing of the ITS-2 rDNA region revealed a single haplotype in all cases. Phylogenetic comparison of P. epiclitum ITS2-rDNA sequences with those from other Paramphistomum, Calicophoron, Gastrothylax and Fischoederius species was performed to assess within and between species variation and validate the use of ITS-2 rDNA as a robust species-specific marker for P. epiclitum identification. This work provides a validated species-specific marker of P. epiclitum and the first report of this parasite species from Pakistan. The results of this study also have implications for the diagnosis and control of rumen flukes in the region and the need for accurate species identification to understand parasite distribution and population genetics.     

Molecular cloning, expression and purification of recombinant soluble mouse endostatin as an anti-angiogenic protein in Escherichia coli

Inhibition of angiogenesis has become a particular interest for treatment of solid tumors. Endostatin, a C-terminal fragment of collagen XVIII, has been reported to exhibit potent inhibitory effect on endothelial cells proliferation, migration and tube formation. In this research, the cDNA library of endostatin was synthesized from mouse liver and inserted into the SacI and SalI enzyme-cutting sites of pUC18 cloning vector. The recombinant vector was transferred into Escherichia coli DH5a and the recombinant clone was selected on LB agar plate plus ampicillin. PCR analysis and DNA sequencing proved the presence of intact endostatin gene in pUC18. The endostatin gene subcloned into pET32a expression vector and the competent bacterial cells of E. coli BL21 were transformed by the vector harboring endostatin gene. In the optimum conditions, expression plasmid was induced with IPTG and recombinant soluble endostatin as a fusion with thioredoxin was purified with Ni–NTA (Ni²⁺-nitrilotriacetate) resin. The results showed that soluble recombinant endostatin as a fusion protein with thioredoxin is a homogenous polypeptide that inhibits angiogenesis (capillary tube formation) in human umbilical vein endothelial cells by 200 ng/ml.     

Long-term administration of rosuvastatin prevents contractile and electrical remodelling of diabetic rat heart

In recent years, many findings have been presented about the potential benefit of statin therapy on diabetes-induced cardiovascular complications. Cardioprotective effects of statins were suggested to be mediated at least in part through inhibition of small GTPases, particularly those of the Rho family. The present study was designed to examine whether rosuvastatin can improve electrical remodeling and contractile dysfunction in type 1 diabetic rat heart via modulation of RhoA pathway. Type 1 diabetes was induced by single dose injection of STZ (50 mg/kg). One week after injection rosuvastatin (10 mg/kg/day) and sham treatment was given for 5 weeks in the diabetic rats, as well as in control groups. Shortening and Ca²⁺ transients were recorded in myocytes loaded with Fura2-AM. Membrane currents and Ca²⁺ transients were measured synchronously via whole-cell patch clamping. In untreated diabetic rats, relaxation of shortening and decay of the matched Ca²⁺ transients were prolonged. Fractional shortening and Ca²⁺ transients were also decreased. Rosuvastatin treatment reversed those changes. I_CaL density did not change in either group but rosuvastatin recovered the loss of sarcoplasmic reticulum Ca²⁺ and Na⁺/Ca²⁺ exchange as evidenced from amplitude and decay of caffeine-induced Ca²⁺ transients, peak I_NCX and calculated sarcoplasmic reticulum Ca²⁺ content. Diabetes-induced attenuation of I_to and I_sus was also reversed, whilst I_K1 was unchanged in diabetes and unaffected by treatment. Rosuvastatin prevented the diabetes-induced increase in RhoA expression. Plasma cholesterol and triglyceride levels were higher in diabetic rats, but rosuvastatin reduced only the latter. In conclusion, HMG-CoA reductase inhibitor rosuvastatin can prevent diabetes-induced electrical and functional remodeling of heart due to inhibition of RhoA signalling rather than reduction of cholesterol level.     

Phylogenetic analysis of coat protein gene of BYDV-MAV strain from wheat

Barley yellow dwarf disease is globally the most important viral disease of wheat. The full-length nucleotide sequence of coat protein (CP) gene of 12 isolates revealed the presence of three distinct clusters. Pakistani isolate of MAV (MAV-PK) has maximum similarity of 99.23% with MAV isolate of Morocco and PAV-Australia following 99.22 and 99.22% with PAV-France. Similar degree of similarity was found in comparison of amino acid sequence. The finding of this study is that MAV-PK has similarity with both MAV-France and PAV-Australia, which is due to the reason that both MAV and PAV belong to the same group and both share maximum nucleotide homology. Low genetic diversity was found not only between MAV isolates but also between MAV and PAV isolates because phylogenetic analysis was done on the CP gene which is highly conserved region in genome of Barley yellow dwarf viruses (BYDVs). Divergence in MAV-PK was due to this recombination which is now most prevalent in Pakistan. MAV-PK has maximum similarity with MAV-Morocco followed by MAV-Sweden and MAV-Cz, which seems to indicate that Pakistani isolate of MAV evolved as the result of recombination between MAV isolates of the USA and PAV isolates of Australia and France. At the same time, recombination of MAV-CZ and MAV-Sweden also occur. This work can be successfully utilised in epidemiological studies of MAV isolate in Pakistan. Further analysis of variation level in these isolates will help scientists to formulate appropriate management strategies like incorporation of BdV 2 gene in wheat against BYDVs.     

Involvement of presenilin holoprotein upregulation in calcium dyshomeostasis of Alzheimer's disease

Mutations in presenilins (PS1 and PS2) account for the vast majority of early onset familial Alzheimer's disease cases. Beside the well investigated role of presenilins as the catalytic unit in γ‐secretase complex, their involvement in regulation of intracellular calcium homeostasis has recently come into more focus of Alzheimer's disease research. Here we report that the overexpression of PS1 full‐length holoprotein forms, in particular familial Alzheimer's disease‐causing forms of PS1, result in significantly attenuated calcium release from thapsigargin‐ and bradykinin‐sensitive stores. Interestingly, treatment of HEK293 cells with γ‐secretase inhibitors also leads to decreased amount of calcium release from endoplasmic reticulum (ER) accompanying elevated PS1 holoprotein levels. Similarly, the knockdown of PEN‐2 which is associated with deficient PS1 endoproteolysis and accumulation of its holoprotein form also leads to decreased ER calcium release. Notably, we detected enhanced PS1 holoprotein levels also in postmortem brains of patients carrying familial Alzheimer's disease PS1 mutations. Taken together, the conditions in which the amount of full length PS1 holoprotein is increased result in reduction of calcium release from ER. Based on these results, we propose that the disturbed ER calcium homeostasis mediated by the elevation of PS1 holoprotein levels may be a contributing factor to the pathogenesis of Alzheimer's disease.