The effect of intrauterine inflammation on mTOR signaling in mouse fetal brain

Fetuses exposed to an inflammatory environment are predisposed to long‐term adverse neurological outcomes. However, the mechanism by which intrauterine inflammation (IUI) is responsible for abnormal fetal brain development is not fully understood. The mechanistic target of rapamycin (mTOR) signaling pathway is closely associated with fetal brain development. We hypothesized that mTOR signaling might be involved in fetal brain injury and malformation when fetuses are exposed to the IUI environment. A well‐established IUI model was utilized by intrauterine injection of lipopolysaccharide (LPS) to explore the effect of IUI on mTOR signaling in mouse fetal brains. We found that microglia activation in LPS fetal brains was increased, as demonstrated by elevated Iba‐1 protein level and immunofluorescence density. LPS fetal brains also showed reduced neuronal cell counts, decreased cell proliferation demonstrated by low Ki67‐positive density, and elevated neuron apoptosis evidenced by high expression of cleaved Caspase 3. Furthermore, we found that mTOR signaling in LPS fetal brains was elevated at 2 hr after LPS treatment, declined at 6 hr and showed overall inhibition at 24 hr. In summary, our study revealed that LPS‐induced IUI leads to increased activation of microglia cells, neuronal damage, and dynamic alterations in mTOR signaling in the mouse fetal brain. Our findings indicate that abnormal changes in mTOR signaling may underlie the development of future neurological complications in offspring exposed to prenatal IUI.     

Cytologic evaluation of prognostic markers in breast carcinoma

To type breast carcinomaon on fine needle aspiration cytology (FNAC) material and correlate the results with histologic typing, to grade breast carcinoma on FNAC material and correlate the findings with Bloom-Richardson histologic grading, and to determine the estrogen receptor (ER) status in cases of breast carcinoma by immunocytochemical (ICC) staining of FNA cytologic material and correlate the findings with ER status, as determined by immunohistochemical (IHC) staining of tissue sections. STUDY DESIGN: Seventy-seven cases of breast carcinoma diagnosed on FNAC formed the basis of this study. Typing was done in all cases on the basis of cytologic features and grading in 62. (Fifteen cases were special types of breast carcinoma). In all cases, ER status was determined by immunostaining of cytologic smears. Results of tumor typing, grading and ER status on cytologic material were compared with the results of histologic typing, grading and immunostaining of histologic material obtained from mastectomy or wide excision specimens. RESULTS: Tumor typing was accurate in 73 of 77 cases (94.8%). Fifteen of 18 cases that were cytologically grade 3 were confirmed on histology, while 3 proved to be grade 2. Of 40 cytologic grade 2 cases, 26 were confirmed on histology, while 14 cases were grade 3. Three of 4 cytologically grade 1 cases were confirmed on histology while 1 was grade 2. The overall accuracy for cytologic grading was 71% (44 of 62 cases). Thirty-seven of 40 ER-positive cases (92.5%) were labeled ER positive on ICC. One case was ER negative on cytology, while in 2 cases the cellularity of the cytologic smear was insufficient to assess ER expression. Thirty-seven cases were negativefor ER on IHC. Nine of these showed ER positivity on ICC, 26 were negative, and 2 had cellularity that was inadequate for assessment of ER. Sensitivity and specificity rates for ER detection on ICC were 97.4% and 74.3%, respectively. CONCLUSION: Tumor typing, grading and evaluation of ER status on FNA C material in breast carcinomas are simple, quick and moderately reliable techniques that compare and correlate favorably with histologic typing, grading and ER status on IHC.     

Bioremediation of cadmium induced renal toxicity in Rattus norvegicus by medicinal plant Catharanthus roseus

Cadmium is the second most hazardous metals with bio-concentration factor (BCF)?>?100 Although WHO permitted cadmium concentration in drinking water is 0.005?mg/L, yet the reality is far above to this limit because of industrial utility of this metal. Oral exposure of cadmium to human results in dreadful symptoms of metabolic disorders especially in liver and kidneys. Endogenous protection could be supported by some exogenous herbal supplement (viz., Catharanthus roseus in this case) to overcome the toxic effects. Present Study has been designed to find out the functional renal changes under the effect of cadmium and Catharanthus roseus in the model organism albino rats. Cadmium significantly (p?>?0.01) increases the level of nitrogenous waste (Urea, BUN, Uric Acid and Creatinin), while decreases the serum protein profile in acute and sub-acute sets. Urea concentration of control ranged from 16.56 to 17.72?mg/dl while that of Group-B and D were 19.84 to 20.87?mg/dl and 17.56 to 17.59?mg/dl respectively. Similarly uric acid concentration ranged in control form 6.98 to 8.01?mg/dl in group-B from 7.58 to 10.25?mg/dl, in Group-D 8.02 to 8.59?mg/dl respectively. Creatinin concentration ranged in control 0.57 to 0.65?mg/dl, in group-B 0.97 to 1.02?mg/dl, in group-D – 0.95 to 0.98?mg/dl respectively.These results might be due to altered filtration rate of kidney because of protein disruption. The studies conclude the efficient nephro-protection offered by Catharanthus roseus extract against Cadmium toxicity.     

Osteoactivin, an anabolic factor that regulates osteoblast differentiation and function

Osteoactivin (OA) is a novel glycoprotein that is highly expressed during osteoblast differentiation. Using Western blot analysis, our data show that OA protein has two isoforms, one is transmembranous and the other is secreted into the conditioned medium of primary osteoblasts cultures. Fractionation of osteoblast cell compartments showed that the mature, glycosylated OA isoform of 115 kDa is found in the membranous fraction. Both OA isoforms (secreted and transmembrane) are found in the cytoplasmic fraction of osteoblasts. Overexpression of EGFP-tagged OA in osteoblasts showed that OA protein accumulates into vesicles for transportation to the cell membrane. We examined OA protein production in primary osteoblast cultures and found that OA is maximally expressed during the third week of culture (last stage of osteoblast differentiation). Glycosylation studies showed that OA isoform of 115 kDa is highly glycosylated. We also showed that retinoic acid (RA) stimulates the mannosylation of OA protein. In contrast, tunicamycin (TM) strongly inhibited N-glycans incorporation into OA protein. The functional role of the secreted OA isoform was revealed when cultures treated with anti-OA antibody, showed decreased osteoblast differentiation compared to untreated control cultures. Gain-of-function in osteoblasts using the pBABE viral system showed that OA overexpression in osteoblast stimulated their differentiation and function. The availability of a naturally occurring mutant mouse with a truncated OA protein provided further evidence that OA is an important factor for terminal osteoblast differentiation and mineralization. Using bone marrow mesenchymal cells derived from OA mutant and wild-type mice and testing their ability to differentiate into osteoblasts showed that differentiation of OA mutant osteoblasts was significantly reduced compared to wild-type osteoblasts. Collectively, our data suggest that OA acts as a positive regulator of osteoblastogenesis.     

Nucleosides and Nucleotides. 125. Synthesis and Biological Evaluation of 2′,3′-Dideoxy-3′-fluoro-2′-methylidene Pyrimidine Nucleosides

Abstract

Reaction of 2′-deoxy-2′-methylidene-5′-O-trityluridine (1) with diethylamino-sulfur trifluoride (DAST) in CH₂Cl₂ resulted in the formation of a mixture of (3′R)-2′,3′-dideoxy-3′-fluoro-2′-methylidene derivative 3 and 2′,3′-didehydro-2′,3′-dideoxy-2′-fluoromethyl derivative 4 (3:4 = 1:1.5) in 65% yield. A similar treatment of 1-(2-deoxy-2-methylidene-5-O-trityl-β-D-threo-pentofuranosyl)uracil (19) with DAST in CH₂Cl₂ afforded (3′S)-2′,3′-dideoxy-3′-fluoro-2′-methylidene derivatives 20 and 4 in 38% and 17% yields respectively. Transformation of the uracil nucleosides 4, 12, and 20 into cytosines followed by deprotection furnished the corresponding cytidine derivatives 29, 18, and 25, respectively. The corresponding thymidine congener 27 was also synthesized in a similar manner. All of the newly synthesized nucleosides were evaluated for their inhibitory activities against HIV and for their antiproliferative activities against L1210 and KB cells.     

Fungal lysis by a soil bacterium fermenting cellulose

Recycling of plant biomass by a community of bacteria and fungi is fundamental to carbon flow in terrestrial ecosystems. Here we report how the plant fermenting, soil bacterium Clostridium phytofermentans enhances growth on cellulose by simultaneously lysing and consuming model fungi from soil. We investigate the mechanism of fungal lysis to show that among the dozens of different glycoside hydrolases C. phytofermentans secretes on cellulose, the most highly expressed enzymes degrade fungi rather than plant substrates. These enzymes, the GH18 Cphy1799 and Cphy1800, synergize to hydrolyse chitin, a main component of the fungal cell wall. Purified enzymes inhibit fungal growth and mutants lacking either GH18 grow normally on cellulose and other plant substrates, but have a reduced ability to hydrolyse chitinous substrates and fungal hyphae. Thus, C. phytofermentans boosts growth on cellulose by lysing fungi with its most highly expressed hydrolases, highlighting the importance of fungal interactions to the ecology of cellulolytic bacteria.     

Cytotoxicity and gene expression profiles of novel synthesized steroid derivatives as chemotherapeutic anti-breast cancer agents

Anti-cancer agents which combine two biologically active compounds in one such as steroidal heterocyclic derivatives attain both hormone and cytotoxic effects on cancer cells. The aim of the present study is to synthesize and evaluate new potential chemotherapeutic anti-breast cancer agents. Several pyridazino-, pyrimido-, quinazolo-, oxirano- and thiazolo-steroid derivatives were synthesized. The structure of the novel steroid derivatives was confirmed using the analytical and spectral data. The most structurally promising of the novel synthesized steroid derivatives, compounds 8, 12, 17, 20, 22c, 24c, 30a and 30b, were investigated individually as anti-breast cancer agents against human breast cancer cells (MCF-7) using sulforhodamine B (SRB) assay. The tested compounds 17, 20, 22c and 8 showed potent broad spectrum cytotoxic activity in vitro after 48 h incubation. Compound 17 (IC(50)=2.5 μM) exhibited more inhibitory influence on MCF-7 growth than the reference drug doxorubicin (Dox) (IC(50)=4.5 μM) after 48 h incubation. Also, the present study showed that all the tested steroid derivatives exhibited significant depletion with various intensities in gene expression of breast cancer related genes (VEGF, CYP19 and hAP-2γ). Noteworthy, compounds 17, 20 and 22c showed the most pronounced effect in this respect.     

Arbuscular mycorrhizal fungi and biochar improves drought tolerance in chickpea

Arbuscular mycorrhizal fungi (AMF) inoculation and biochar amendment has been reported to improve growth of several crop plants however their role in stress amelioration individually as well as in combination has not been worked out. This experiment was conducted to evaluate the application of AMF and biochar on the performance of chickpea under drought stress. The treatments included the individual as well as combined treatment of AMF and biochar to drought stressed and normal chickpea plants. Plants inoculation improved growth in terms of shoot and root length, leaf area and number of branches which was observed to show a steep decline due to drought stress. Drought declined the AMF colonization potential though biochar amendment ameliorated the negative effects of drought significantly by improving the spore population, number of mycelium, vesicle and arbuscules and the percentage of colonization as well. Increased chlorophyll synthesis in biochar and AMF treated plants was obvious, which lead to significant enhancement in the net photosynthetic efficiency. Drought stress also declined the relative water content (RWC) and membrane stability index (MSI), while treatment of biochar and AMF either individually or in combination mitigated the deleterious effects to considerable extent and caused a significant enhancement in RWC and MSI under normal conditions. Amendments with biochar and AMF inoculation increased the nitrogen fixation attributes including the number and weight of nodules, leghemoglobin content and activity of nitrate reductase enzyme leading to greater uptake and assimilation of nitrogen in them when compared to drought stressed plants. Drought stressed chickpea plants exhibited considerable reduction in uptake of nitrogen and phosphorous which was ameliorated by biochar and AMF treatments. It could be suggested that increase in growth and physiological attributes in chickpea due to biochar amendments and AMF inoculation under drought stress were plausibly due to their involvement in nitrogen and phosphorous uptake, chlorophyll synthesis and photosynthesis.