Characterization of cross-bridge elasticity and kinetics of cross-bridge cycling during force development in single smooth muscle cells

Force development in smooth muscle, as in skeletal muscle, is believed to reflect recruitment of force-generating myosin cross-bridges. However, little is known about the events underlying cross-bridge recruitment as the muscle cell approaches peak isometric force and then enters a period of tension maintenance. In the present studies on single smooth muscle cells isolated from the toad (Bufo marinus) stomach muscularis, active muscle stiffness, calculated from the force response to small sinusoidal length changes (0.5% cell length, 250 Hz), was utilized to estimate the relative number of attached cross-bridges. By comparing stiffness during initial force development to stiffness during force redevelopment immediately after a quick release imposed at peak force, we propose that the instantaneous active stiffness of the cell reflects both a linearly elastic cross-bridge element having 1.5 times the compliance of the cross-bridge in frog skeletal muscle and a series elastic component having an exponential length-force relationship. At the onset of force development, the ratio of stiffness to force was 2.5 times greater than at peak isometric force. These data suggest that, upon activation, cross-bridges attach in at least two states (i.e., low-force-producing and high-force-producing) and redistribute to a steady state distribution at peak isometric force. The possibility that the cross-bridge cycling rate was modulated with time was also investigated by analyzing the time course of tension recovery to small, rapid step length changes (0.5% cell length in 2.5 ms) imposed during initial force development, at peak force, and after 15 s of tension maintenance. The rate of tension recovery slowed continuously throughout force development following activation and slowed further as force was maintained. Our results suggest that the kinetics of force production in smooth muscle may involve a redistribution of cross-bridge populations between two attached states and that the average cycling rate of these cross-bridges becomes slower with time during contraction.     

Release of poly a(+) messenger RNA from rat liver rough microsomes upon disassembly of bound polysomes

Several procedures were used to disassemble rat liver rough microsomes (RM) into ribosomal subunits, mRNA, and ribosome-stripped membrane vesicles in order to examine the nature of the association between the mRNA of bound polysomes and the microsomal membranes. The fate of the mRNA molecules after ribosome release was determined by measuring the amount of pulse-labeled microsomal RNA in each fraction which was retained by oligo-dT cellulose or by measuring the poly A content by hybridization to radioactive poly U. It was found that ribosomal subunits and mRNA were simultaneously released from the microsomal membranes when the ribosomes were detached by: (a) treatment with puromycin in a high salt medium containing Mg++, (b) resuspension in a high salt medium lacking Mg++, and (c) chelation of Mg++ by EDTA or pyrophosphate. Poly A-containing mRNA fragments were extensively released from RM subjected to a mild treatment with pancreatic RNase in a medium of low ionic strength. This indicates that the 3' end of the mRNA is exposed on the outer microsomal surface and is not directly bound to the membranes. Poly A segments of bound mRNA were also accessible to [(3)H] poly U for in situ hybridization in glutaraldehyde-fixed RM. Rats were treated with drugs which inhibit translation after formation of the first peptide bonds or interfere with the initiation of protein synthesis. After these treatments inactive monomeric ribosomes, as well as ribosomes bearing mRNA, remained associated with their binding sites in microsomes prepared in media of low ionic strength. However, because there were no linkages provided by nascent chains, ribosomes, and mRNA, molecules were released from the microsomal membranes without the need of puromycin, by treatment with a high salt buffer containing Mg++. Thus, both in vivo and in vitro observations are consistent with a model in which mRNA does not contribute significantly to the maintenance of the interaction between bound polysomes and endoplasmic reticulum membranes in rat liver hepatocytes.     

Biosynthesis of lysosomal hydrolases: their synthesis in bound polysomes and the role of co- and post-translational processing in determining their subcellular distribution

By in vitro translation of mRNA’s isolated from free and membrane-bound polysomes, direct evidence was obtained for the synthesis of two lysosomal hydrolases, β-glucuronidase of the rat preputial gland and cathespin D of mouse spleen, on polysomes bound to rough endoplasmic reticulum (ER) membranes. When the mRNA’s for these two proteins were translated in the presence of microsomal membranes, the in vitro synthesized polypeptides were cotranslationally glycosylated and transferred into the microsomal lumen. Polypeptides synthesized in the absence of microsomal membranes were approximately 2,000 daltons larger than the respective unglycosylated microsomal polypeptides found after short times of labeling in cultured rat liver cells treated with tunicamycin. This strongly suggests that nascent chains of the lysosomal enzymes bear transient amino terminal signals which determine synthesis on bound polysomes and are removed during the cotranslational insertion of the polypeptides into the ER membranes. In the line of cultured rat liver cells used for this work, newly synthesized lysosomal hydrolases showed a dual destination; approximately 60 percent of the microsomal polypeptides detected after short times of labeling were subsequently processed proteolytically to lower molecular weight forms characteristic of the mature enzymes. The remainder was secreted from the cells without further proteolytic processing. As previously observed by other investigations in cultured fibroblasts (A. Gonzalez-Noriega, J.H. Grubbs, V. Talkad, and W.S. Sly, 1980, J Cell Biol. 85: 839-852; A. Hasilik and E.F. Neufeld, 1980, J. Biol. Chem., 255:4937-4945.) the lysosomotropic amine chloroquine prevented the proteolytic maturation of newly synthesized hydrolases and enhanced their section. In addition, unglycosylated hydrolases synthesized in cells treated with tunicamycin were exclusively exported from the cells without undergoing proteolytic processing. These results support the notions that modified sugar residues serve as sorting out signals which address the hydrolases to their lysosomal destination and that final proteolytic cleavage of hydrolase precursors take place within lysosome itself. Structural differences in the carbohydrate chains of intracellular and secreted precursors of cathespin D were detected from their differential sensitivity to digestion with endoglycosidases H and D. These observations suggest that the hydrolases exported into the medium follow the normal secretory route and that some of their oligosaccharides are subject to modifications known to affect many secretory glycoproteins during their passage through the Golgi apparatus.     

Melanin–Perovskite Composites for Photothermal Conversion

Biomacromolecular pigments, such as melanin, play an essential role in the survival of all living beings. Melanin absorbs sunlight and transforms it into heat, which is crucial for avoiding damage to skin cells. Light absorption produces excited electrons, which could either fall back to ground states by releasing the heat (photothermal effect) and/or light (photoluminescence), or stay at higher energy levels within its lifetime period, which can be captured through external electronic circuitry (photovoltaic effect). In this study, it is demonstrated that the combination of melanin with halide perovskite light absorber in the form of a composite exhibits high absorbance from the UV to NIR region in the solar spectrum. And the composite displays significantly reduced photoluminescence and minimized density of residual excited states (verified by photovoltaic measurement) owing to the significantly enhanced nonradiant quenching by the melanin. As a result, the composite shows an ultrahigh solar‐thermal quantum yield of 99.56% and solar‐thermal conversion efficiency of ≈81% under one‐sun illumination (AM1.5), which is superior to typical carbon materials such as graphene (≈70%). By coating the photothermal composite film on the hot‐side of thermoelectric devices, a 7000% increase in output power as compared to the blank device under illumination is observed.     

Mechanism of Acetic Acid Gustatory Repulsion in Drosophila

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Infestation of people with lice in Kathmandu and Pokhara, Nepal

The prevalence of infestation with head lice and body lice, Pediculus spp. (Phthiraptera: Pediculidae) and pubic (crab) lice Pthirus pubis (L.) (Phthiraptera: Pthiridae), was recorded from 484 people in Nepal. The prevalence of head lice varied from 16% in a sample of people aged 10-39 years of age, to 59% in street children. Simultaneous infestations with head and body lice (double infestations) varied from 18% in slum children to 59% in street children.     

Losing the desire: selection can promote obligate asexuality

Whilst parthenogenesis has evolved multiple times from sexual invertebrate and vertebrate lineages, the drivers and consequences of the sex-asex transition remain mostly uncertain. A model by Stouthamer et al. recently published in BMC Evolutionary Biology shows a pathway by which obligate asexuality could be selected for following endosymbiont infection.     

Role of the C8 gem-dimethyl group of bryostatin 1 on its unique pattern of biological activity

The role of the C(8) gem-dimethyl group in the A-ring of bryostatin 1 has been examined through chemical synthesis and biological evaluation of a new analogue. Assays for biological function using U937, K562, and MV4-11 cells as well as the profiles for downregulation of PKC isozymes revealed that the presence of this group is not a critical determinant for the unique pattern of biological activity of bryostatin.     

High Risk of ART Non-Adherence and Delay of ART Initiation among HIV Positive Double Orphans in Kigali, Rwanda

Background

To reduce HIV/AIDS related mortality of children, adherence to antiretroviral treatment (ART) is critical in the treatment of HIV positive children. However, little is known about the association between ART adherence and different orphan status. The aims of this study were to assess the ART adherence and identify whether different orphan status was associated with the child’s adherence.

Methods

A total of 717 HIV positive children and the same number of caregivers participated in this cross-sectional study. Children’s adherence rate was measured using a pill count method and those who took 85% or more of the prescribed doses were defined as adherent. To collect data about adherence related factors, we also interviewed caregivers using a structured questionnaire.

Results

Of all children (N = 717), participants from each orphan category (double orphan, maternal orphan, paternal orphan, non-orphan) were 346, 89, 169, and 113, respectively. ART non-adherence rate of each orphan category was 59.3%, 44.9%, 46.7%, and 49.7%, respectively. The multivariate analysis indicated that maternal orphans (AOR 0.31, 95% CI 0.12–0.80), paternal orphans (AOR 0.35, 95% CI 0.14–0.89), and non-orphans (AOR 0.45, 95% CI 0.21–0.99) were less likely to be non-adherent compared to double orphans. Double orphans who had a sibling as a caregiver were more likely to be non-adherent. The first mean CD4 count prior to initiating treatment was 520, 601, 599, and 844 (cells/ml), respectively (p<0.001). Their mean age at sero-status detection was 5.9, 5.3, 4.8, and 3.9 (year old), respectively (p<0.001).

Conclusions

Double orphans were at highest risk of ART non-adherence and especially those who had a sibling as a caregiver had high risk. They were also in danger of initiating ART at an older age and at a later stage of HIV/AIDS compared with other orphan categories. Double orphans need more attention to the promote child’s adherence to ART.