Studies on bone marrow histogenesis: Morphometric and autoradiographic studies of regenerating marrow stroma in extramedullary autoimplants and after evacuation of marrow cavity

Summary The marrow cavity of the rat tibia was mechanically evacuated and autoimplanted to the subcutaneous tissue. The regenerative process which restored the integrity of marrow stroma and hemopoiesis, was morphometrically evaluated in whole mount of tibia. Following evacuation, the clot filled the cavity. The granulation tissue then appeared and expanded, penetrating and replacing the clot. The fibroblasts of the granulation tissue differentiated into osteoblasts forming osteoid bone. Within its interstices, the primordial marrow consisting of loose connective tissue and vascular sinuses appeared and hemopoiesis resumed. Expansion of hemopoiesis resulted in the resorption of bone and within three weeks the tibial cavity was restored to the pre-evacuation state.Autoradiography indicated that the labeling index was initially high in fibroblasts and osteoblasts but was subsequently reduced while it increased in osteocytes, cells of Haversian canals, stromal and hemopoietic cells of marrow. The finding is in disagreement with the view that the regenerative process originates from the Haversian canal. When the label was introduced on day 4 post-operatively, it subsequently appeared in osteocytes, cells of Haversian canal, stromal elements of the marrow, but not in the hemopoietic cells. This indicates complete dissociation of marrow stroma and hemopoietic stem cell.Supported by NASA Contract NSG 9061. Mehdi Tavassoli is the recipient of a CRD Award AM-70551     

Prostaglandin I2 stimulation of granulosa cell cyclic AMP production

The ability of prostaglandin I₂ (PGI₂) to stimulate cyclic AMP production by granulosa cells, isolated from intact immature rats, has been demonstrated in vitro. The minimal effective dose was 15 ng/ml, which was comparable to the minimal effective dose for PGE₂. However, a concentration of 15 μg/ml PGI₂ was required to stimulate cyclic AMP production maximally, compared to a concentration of 1 μg/ml PGE₂, which produced the maximum response. It therefore appears that PGI₂ is not more effective than PGE₂ in stimulating cyclic AMP production in granulosa cells, and is possibly less effective. Submaximal concentrations of PGI₂ appeared to be able to modify the stimulation of cyclic AMP production by follicle- stimulating hormone (FSH), but whether or not PGI₂ plays any role in follicular function remains to be established.     

Characterization of Salmonella spp. isolated from patients below 3 years old with acute diarrhoea

Fifteen strains of Salmonella were isolated from children with clinically diagnosed diarrhoea aged below 3 years old, who had been admitted to K7 ward, Pediatric Institute, Kuala Lumpur Hospital. The isolates were tested for their susceptibility to a range of antimicrobial agents, and typed by serological tests and randomly amplified polymorphic DNA (RAPD) fingerprinting. All the strains had a similar pattern of antimicrobial susceptibility, where they were susceptible to a wide range of antimicrobial agents. The serological test has typed them into three serovars, which were identified as Salmonella enterica ser. Akanji, Salmonella enterica ser. Hindmarch and Salmonella enterica ser. Richmond. In contrast, the RAPD fingerprinting classed them into two major clusters, cluster 1 consisting of 12 strains of Salmonella and cluster 2 consisting of three strains of Salmonella.     

Tsetse blood-meal sources,endosymbionts and trypanosome-associations in the Maasai Mara National Reserve,a wildlife-human-livestock interface

African trypanosomiasis (AT) is a neglected disease of both humans and animals caused by Trypanosoma parasites, which are transmitted by obligate hematophagous tsetse flies (Glossina spp.). Knowledge on tsetse fly vertebrate hosts and the influence of tsetse endosymbionts on trypanosome presence, especially in wildlife-human-livestock interfaces, is limited. We identified tsetse species, their blood-meal sources, and correlations between endosymbionts and trypanosome presence in tsetse flies from the trypanosome-endemic Maasai Mara National Reserve (MMNR) in Kenya. Among 1167 tsetse flies (1136 Glossina pallidipes, 31 Glossina swynnertoni) collected from 10 sampling sites, 28 (2.4%) were positive by PCR for trypanosome DNA, most (17/28) being of Trypanosoma vivax species. Blood-meal analyses based on high-resolution melting analysis of vertebrate cytochrome c oxidase 1 and cytochrome b gene PCR products (n = 354) identified humans as the most common vertebrate host (37%), followed by hippopotamus (29.1%), African buffalo (26.3%), elephant (3.39%), and giraffe (0.84%). Flies positive for trypanosome DNA had fed on hippopotamus and buffalo. Tsetse flies were more likely to be positive for trypanosomes if they had the Sodalis glossinidius endosymbiont (P = 0.0002). These findings point to complex interactions of tsetse flies with trypanosomes, endosymbionts, and diverse vertebrate hosts in wildlife ecosystems such as in the MMNR, which should be considered in control programs. These interactions may contribute to the maintenance of tsetse populations and/or persistent circulation of African trypanosomes. Although the African buffalo is a key reservoir of AT, the higher proportion of hippopotamus blood-meals in flies with trypanosome DNA indicates that other wildlife species may be important in AT transmission. No trypanosomes associated with human disease were identified, but the high proportion of human blood-meals identified are indicative of human African trypanosomiasis risk. Our results add to existing data suggesting that Sodalis endosymbionts are associated with increased trypanosome presence in tsetse flies.     

Proline-Functionalized Graphene Oxide Nanoparticles (GO–Pro NPs) Mitigate Salt-Induced Adverse Effects on Morpho-Physiological Traits and Essential Oils Constituents in Moldavian Balm (Dracocephalum moldavica L.)

Many methodologies have been established to lessen negative impacts of salinity on plants. Of those methodologies, nanoparticles (NPs) application has achieved great importance thanks to their unique physico-chemical properties. Consequently, formerly respecting encouraging impacts of graphene oxide (GO) and proline (Pro) on different plant processes under non-stress and stress conditions, proline-functionalized graphene oxide nanoparticles “GO–Pro NPs” were synthesized and characterized. Graphite powder, as starting material, was used to synthesize GO using modified Hummers method followed by functionalization of its surface by proline in basic media. Afterward, GO–Pro NPs, GO and Pro, each at 0, 50 and 100 mg L^?1 concentrations with three replications, were applied on Moldavian balm (Dracocephalum moldavica L.) plants to assay their effects under non-stress (0 mM) and salt stress (50 and 100 mM) conditions. GO–Pro NPs and Pro effectively alleviated negative effects of salinity through increasing morphological parameters, photosynthetic pigments, chlorophyll fluorescence parameters, chlorophyll index (SPAD), and membrane stability index (MSI) and decreasing hydrogen peroxide and malondialdehyde, as well. Also application of GO–Pro NPs enhanced proline, antioxidant enzymes activities, and most dominant constituents of essential oil. The highest MSI (48.87%) and proline content (15.36 µM g^?1 FW) were observed in plant treated with GO–Pro NPs (50 mg L^?1) under 100 mM NaCl salinity stress. The GO–Pro NPs treatment at lower dose (50 mg L^?1) could be introduced as the best preservative treatment for Moldavian balm under salt stress. GO application mostly had no effect on the measured parameters announcing it as carrier for Pro to enhance its efficiency. In conclusion, GO–Pro NPs application could promote Moldavian balm performance and essential oil under salinity presenting GO–Pro NPs as new treatment against stress conditions.

     

Structural requirements within the lipoyl domain for the Ca2+-dependent binding and activation of pyruvate dehydrogenase phosphatase isoform 1 or its catalytic subunit

The inner lipoyl domain (L2) of the dihydrolipoyl acetyltransferase (E2) 60-mer forms a Ca(2+)-dependent complex with the pyruvate dehydrogenase phosphatase 1 (PDP1) or its catalytic subunit, PDP1c, in facilitating large enhancements of the activities of PDP1 (10-fold) or PDP1c (6-fold). L2 binding to PDP1 or PDP1c requires the lipoyl-lysine prosthetic group and specificity residues that distinguish L2 from the other lipoyl domains (L1 in E2 and L3 in the E3-binding component). The L2-surface structure contributing to binding was mapped by comparing the capacities of well folded mutant or lipoyl analog-substituted L2 domains to interfere with E2 activation by competitively binding to PDP1 or PDP1c. Our results reveal the critical importance of a regional set of residues near the lipoyl group and of the octanoyl but not the dithiolane ring structure of the lipoyl group. At the other end of the lipoyl domain, substitution of Glu(182) by alanine or glutamine removed L2 binding to PDP1 or PDP1c, and these substitutions for the neighboring Glu(179) also greatly hindered complex formation (E179A > E179Q). Among 11 substitutions in L2 at sites of major surface residue differences between the L1 and L2 domains, only the conversion of Val-Gln(181) located between the critical Glu(179) and Glu(182) to the aligned Ser-Leu sequence of the L1 domain greatly reduced L2 binding. Certain modified L2 altered E2 activation of PDP1 differently than PDP1c, supporting significant impact of the regulatory PDP1r subunit on PDP1 binding to L2. Our results indicate hydrophobic binding via the extended aliphatic structure of the lipoyl group and required adjacent L2 structure anchor PDP1 by acting in concert with an acidic cluster at the other end of the domain.