PBX,MEIS, and IGF-I are potential mediators of retinoic acid-induced proximodistal limb reduction defects

BACKGROUND: Phocomelia, which is primarily due to a disruption in the proximodistal axis, is found in virtually all mouse embryos exposed to high doses of retinoic acid (RA) on 11 days post coitum (dpc). METHODS: To identify genes that potentially mediate the effects of retinoic acid (RA) on limb development, we have examined the expression of 9,000 clones from the IMAGE consortium by microarray analysis of RNA isolated from 11 dpc mouse forelimbs exposed to RA or vehicle for 6 hr. Eight genes that demonstrated altered expression were chosen for further study of their mRNA levels using RT-PCR. Protein levels were determined by Western blot analysis. RESULTS: Of the 9,000 genes examined in the microarray, approximately 111 demonstrated altered expression (33 known genes and 78 ESTs). Of the eight known genes chosen for further study using RT-PCR, four mRNAs (PBX1a, PBX1b, IGF-Ia, and IGF-Ib) demonstrated consistent elevation ( approximately 3-fold) in their levels after RA treatment in both the forelimbs and hindlimbs as early as 3 hr after RA treatment. In addition to the two PBX1 isoforms, the mRNA level of the other two subtypes (PBX2 and PBX3) and the level of PBX1/2/3 protein were also found to be elevated in limb buds after RA treatment. Finally, we examined the expression of MEIS1, MEIS2, and MEIS3 because these proteins are necessary for PBX nuclear localization. The mRNA level of all three subtypes of MEIS were elevated approximately three- to four-fold in both the forelimbs and hindlimbs after RA treatment. CONCLUSIONS: Because both PBX and MEIS (and their orthologs) are believed to be involved in the control of proximodistal axis formation in mouse and fly limbs and IGFs in the development of limbs, we suggest that increases in PBX, MEIS and IGF-1 mRNA levels may contribute to proximodistal limb reduction defects caused by teratogenic doses of RA.     

Effect of different oxidation states of chromium in causing chromosome alterations in cultured CHO cells

Four chromium salts with different oxidation states were tested for their influence in causing chromosome aberrations and sister-chromatid exchange in Chinese hamster ovary cellsin vitro. Cell cultures were treated with CrO₃, K₂Cr₂O₇, CrCl₂ and Cr(NO₃)₃.9H₂O at concentrations of 10^–7, 10^–6, 10^–5 and 10^–4 M for the aberration assay, and 10^–8, 10^–7, 10^–6 and 10^–5 M for the sister-chromatid exchange assay. It was noticed that Cr (VI) compounds-CrO₃ and K₂Cr₂O₇-considerably enhanced the frequencies of aberrations and sister-chromatid exchanges compared to the control cultures. CrCl₂ and Cr(NO₃)₃.9H₂O–Cr (II) and Cr (III) respectively-caused a slight increase in sister-chromatid exchange rates, but the frequencies of aberrations were almost unchanged compared to the controls. These investigations indicate a definite link between the metals and changes produced in the mammalian chromosomes, reaffirming the evidence of carcinogenic potential of Cr (VI) observed by other investigators.Abbreviations BrdU 5-bromo-2-deoxyuridine - CHO Chinese hamster ovary - SCE sister-chromatid exchange     

Biochemical characterization of rat intestine development using high-resolution magic-angle-spinning 1H NMR spectroscopy and multivariate data analysis

We report details of metabolic profiles for small intestinal samples obtained using high-resolution magic-angle-spinning (HRMAS) (1)H NMR spectroscopy. Intact samples of jejunum and ileum from male Long Evans rats were analyzed on a 600 MHz spectrometer using standard one and two-dimensional (1)H NMR spectroscopic pulse sequences. The metabolic profiles of ileum and jejunum predominantly comprised a number of amino acids, lipids, glycerophosphocholine (GPC), choline, creatine, and ethanol, a number of carboxylic acids including acetate and lactate, and nucleoside bases including cytosine, isocytosine, and uracil. Principal component analysis (PCA) was applied to these NMR data to characterize the biochemical differences between jejunum and ileum tissues. Compared with ileum, jejunum contained higher levels of lipids, GPC, choline, lactate and creatinine, but lower levels of amino acids and acetate. In addition, the age dependence of the biochemical composition of intestinal tissues from young rats (15, 36 days and 3-4 months old) was studied. In general, levels of lipids, lactate, taurine and creatinine were positively correlated with age while amino acids and GPC decreased in the older age group. This study will provide a metabolic reference for further studies assessing the metabolic consequences of nutrition, stress and gut microbiota on intestinal composition.     

Effectiveness Study of Paromomycin IM Injection (PMIM) for the Treatment of Visceral Leishmaniasis (VL) in Bangladesh

Background

This study was conducted in Bangladeshi patients in an outpatient setting to support registration of Paromomycin Intramuscular Injection (PMIM) as a low-cost treatment option in Bangladesh.

Methodology

This Phase IIIb, open-label, multi-center, single-arm trial assessed the efficacy and safety of PMIM administered at 11 mg/kg (paromomycin base) intramuscularly once daily for 21 consecutive days to children and adults with VL in a rural outpatient setting in Bangladesh. Patients ≥5 and ≤55 years were eligible if they had signs and symptoms of VL (intermittent fever, weight loss/decreased appetite, and enlarged spleen), positive rK39 test, and were living in VL-endemic areas. Compliance was the percentage of enrolled patients who received 21 daily injections over no more than 22 days. Efficacy was evaluated by initial clinical response, defined as resolution of fever and reduction of splenomegaly at end of treatment, and final clinical response, defined as the absence of new clinical signs and symptoms of VL 6 months after end of treatment. Safety was assessed by evaluation of adverse events.

Principal Findings

A total of 120 subjects (49% pediatric) were enrolled. Treatment compliance was 98.3%. Initial clinical response in the Intent-to-Treat population was 98.3%, and final clinical response 6 months after end of treatment was 94.2%. Of the 119 subjects who received ≥1 dose of PMIM, 28.6% reported at least one adverse event. Injection site pain was the most commonly reported adverse event. Reversible renal impairment and/or hearing loss were reported in 2 subjects.

Conclusions/Significance

PMIM was an effective and safe treatment for VL in Bangladesh. The short treatment duration and lower cost of PMIM compared with other treatment options may make this drug a preferred treatment to be investigated as part of a combination therapy regimen. This study supports the registration of PMIM for use in government health facilities in Bangladesh.

Trial Registration

ClinicalTrials.gov identifier: NCT01328457     

Marginal bone loss around axial and straight implants supported with prefabricated SFI-Bar with mandibular overdentures

To assess the role of prefabricated SFI-Bar in peri-implant bone loss around immediately axially loaded and straight implants. This study comprised of 40 complete denture wearer patients who received two axially parallel implants connected by SFI-Bars in group I and two 15° mesially tilted implants connected by SFI-Bars in group II. Peri- implant bone loss (PiBL) was measured at 1 year, 2 years and 3 years. The mean PiBL at 1 year in group I was 0.21 mm and I group II was 0.22, at 2 years in group I was 0.26 mm and in group II was 0.23 mm and at 3 years, in group I was 0.29 mm and in group II was 0.34 mm. The difference was significant at 3 years (P< 0.05). The mean mesial PIBL at 1 year in group I was 0.18 mm, in group II was 0.20 mm, at 2 years in group I was 0.19 mm and in group II was 0.07 mm and at 3 years, in group I was 0.25 mm and in group II was 0.29 mm. The difference found to be significant in each time duration in both groups (P< 0.05).The mean distal PIBL at 1 year in group I was 0.23 mm, in group II was 0.22 mm, at 2 years in group I was 0.33 mm and in group II was 0.39 mm and at 3 years, in group I was 0.34 mm and in group II was 0.39 mm. The difference found to be significant at 2 and 3 years in both groups (P< 0.05). Authors found that mandibular overdentures retained with Prefabricated SFI-Bar with axial and straight inserted implants may be useful in patients with reduced bone height.     

Global metabolic profiling analysis on human urine by UPLC-TOFMS: issues and method validation in nutritional metabolomics

Optimisation and method validation was assessed here for metabolic profiling analysis of urine samples using UPLC-TOFMS. A longer run time of 31 min revealed greater reproducibility, and the higher number of variables was identified as compared to shortened run times (10 and 26 min). We have also implemented two QC urine samples enabling the assessment of the quality and reproducibility of the data generated during the whole analytical workflow (retention time drift, mass precision and fluctuation of the ion responses over time). Based on the QC data, suitable standards for ensuring consistent analytical results for metabolomics applications using the UPLC-MS techniques are recommended.     

Systemic multicompartmental effects of the gut microbiome on mouse metabolic phenotypes

To characterize the impact of gut microbiota on host metabolism, we investigated the multicompartmental metabolic profiles of a conventional mouse strain (C3H/HeJ) (n=5) and its germ‐free (GF) equivalent (n=5). We confirm that the microbiome strongly impacts on the metabolism of bile acids through the enterohepatic cycle and gut metabolism (higher levels of phosphocholine and glycine in GF liver and marked higher levels of bile acids in three gut compartments). Furthermore we demonstrate that (1) well‐defined metabolic differences exist in all examined compartments between the metabotypes of GF and conventional mice: bacterial co‐metabolic products such as hippurate (urine) and 5‐aminovalerate (colon epithelium) were found at reduced concentrations, whereas raffinose was only detected in GF colonic profiles. (2) The microbiome also influences kidney homeostasis with elevated levels of key cell volume regulators (betaine, choline, myo‐inositol and so on) observed in GF kidneys. (3) Gut microbiota modulate metabotype expression at both local (gut) and global (biofluids, kidney, liver) system levels and hence influence the responses to a variety of dietary modulation and drug exposures relevant to personalized health‐care investigations.