Clathrin-dependent entry and vesicle-mediated exocytosis define insulin transcytosis across microvascular endothelial cells

Transport of insulin across the microvasculature is necessary to reach its target organs (e.g., adipose and muscle tissues) and is rate limiting in insulin action. Morphological evidence suggests that insulin enters endothelial cells of the microvasculature, and studies with large vessel–derived endothelial cells show insulin uptake; however, little is known about the actual transcytosis of insulin and how this occurs in the relevant microvascular endothelial cells. We report an approach to study insulin transcytosis across individual, primary human adipose microvascular endothelial cells (HAMECs), involving insulin uptake followed by vesicle-mediated exocytosis visualized by total internal reflection fluorescence microscopy. In this setting, fluorophore-conjugated insulin exocytosis depended on its initial binding and uptake, which was saturable and much greater than in muscle cells. Unlike its degradation within muscle cells, insulin was stable within HAMECs and escaped lysosomal colocalization. Insulin transcytosis required dynamin but was unaffected by caveolin-1 knockdown or cholesterol depletion. Instead, insulin transcytosis was significantly inhibited by the clathrin-mediated endocytosis inhibitor Pitstop 2 or siRNA-mediated clathrin depletion. Accordingly, insulin internalized for 1 min in HAMECs colocalized with clathrin far more than with caveolin-1. This study constitutes the first evidence of vesicle-mediated insulin transcytosis and highlights that its initial uptake is clathrin dependent and caveolae independent.     

Of PCOS Symptoms,Hirsutism Has the Most Significant Impact on the Quality of Life of Iranian Women

Introduction

Polycystic ovary syndrome is a common endocrine disorder affecting women both physically and psychologically and can lead to a poor quality of life compared to their normal counterparts. The aim of the present study was to assess the impact of various clinical features of polycystic ovary syndrome on the health-related quality of life of Iranian women diagnosed with this syndrome.

Materials and Methods

A total of 796 women diagnosed with polycystic ovary syndrome, aged 15–49 years, completed the questionnaires, interviews, and medical assessments required for this study. A reliable and validated Persian version of the health-related quality of life questionnaire for polycystic ovary syndrome patients was used. Linear regression models were used to assess the association between the symptoms of polycystic ovary syndrome and health-related quality of life.

Results

The mean age of participants was 28.02 years. 35.4% of the subjects were classified as overweight or obese. Hirsutism, was reported to have the strongest impact on the patients’ health-related quality of life, followed in descending order by body mass index, irregular menses and infertility. The relative level of hirsutism was directly proportional to decrease in health-related quality of life score (p<0.001).

Conclusions

The results of the study found that hirsutism had the strongest impact on the health-related quality of life measures in Iranian women diagnosed with polycystic ovary syndrome. Health care officials need to evaluate in depth the effect of each clinical feature of polycystic ovary syndrome separately and design management strategies, keeping in mind the psychological and physical manifestations.     

Peach Water Relations,Gas Exchange,Growth and Shoot Mortality under Water Deficit in Semi-Arid Weather Conditions

In this study the sensitivity of peach tree (Prunus persica L.) to three water stress levels from mid-pit hardening until harvest was assessed. Seasonal patterns of shoot and fruit growth, gas exchange (leaf photosynthesis, stomatal conductance and transpiration) as well as carbon (C) storage/mobilization were evaluated in relation to plant water status. A simple C balance model was also developed to investigate sink-source relationship in relation to plant water status at the tree level. The C source was estimated through the leaf area dynamics and leaf photosynthesis rate along the season. The C sink was estimated for maintenance respiration and growth of shoots and fruits. Water stress significantly reduced gas exchange, and fruit, and shoot growth, but increased fruit dry matter concentration. Growth was more affected by water deficit than photosynthesis, and shoot growth was more sensitive to water deficit than fruit growth. Reduction of shoot growth was associated with a decrease of shoot elongation, emergence, and high shoot mortality. Water scarcity affected tree C assimilation due to two interacting factors: (i) reduction in leaf photosynthesis (-23% and -50% under moderate (MS) and severe (SS) water stress compared to low (LS) stress during growth season) and (ii) reduction in total leaf area (-57% and -79% under MS and SS compared to LS at harvest). Our field data analysis suggested a Ψ_stem threshold of -1.5 MPa below which daily net C gain became negative, i.e. C assimilation became lower than C needed for respiration and growth. Negative C balance under MS and SS associated with decline of trunk carbohydrate reserves – may have led to drought-induced vegetative mortality.     

Differential Gene Expression Reflects Morphological Characteristics and Physiological Processes in Rice Immunity against Blast Pathogen Magnaporthe oryzae

The rice blast fungus Magnaporthe oryzae is a serious pathogen that jeopardises the world’s most important food-security crop. Ten common Malaysian rice varieties were examined for their morphological, physiological and genomic responses to this rice blast pathogen. qPCR quantification was used to assess the growth of the pathogen population in resistant and susceptible rice varieties. The chlorophyll content and photosynthesis were also measured to further understand the disruptive effects that M. oryzae has on infected plants of these varieties. Real-time PCR was used to explore the differential expression of eight blast resistance genes among the ten local varieties. Blast disease has destructive effects on the growth of rice, and the findings of our study provide evidence that the Pikh, Pi9, Pi21, and Osw45 genes are involved in defence responses in the leaves of Malaysian rice at 31 h after inoculation with M. oryzae pathotype P7.2. Both the chlorophyll content and photosynthesis were reduced, but the levels of Pikh gene expression remained constant in susceptible varieties, with a developed pathogen population and mild or severe symptoms. The Pi9, Pi21, and Osw45 genes, however, were simultaneously upregulated in infected rice plants. Therefore, the presence of the Pikh, Pi9, Pi21, and Osw45 genes in the germplasm is useful for improving the resistance of rice varieties.     

Overexpression of reactive oxygen species modulator 1 is associated with advanced grades of bladder cancer

Molecular Biology Reports - Reactive Oxygen Species Modulator 1 (ROMO1) plays a pivotal role in the regulation of mitochondrial structure integrity, and the production of reactive oxygen species...     

Synthesis of folic acid functionalized terbium‐doped dendritic fibrous nano‐silica and Interaction with HEK 293 normal,MDA breast cancer and HT 29 colon cancer cells

A novel folic acid functionalized terbium‐doped dendritic fibrous nanoparticle (Tb@KCC‐1‐NH₂‐FA) with high surface area was synthesized using a novel hydrothermal protocol. In the present work, we report the fluorescent Tb‐doted nanomaterial with emission wavelength at 497 nm which confirms the formation of Tb@KCC‐1‐NH₂‐FA. Synthesized nanoparticles were investigated through transmission electron microscope, field emission scanning electron Microscopy, Fourier transform infrared spectra, Brunauer‐Emmett‐Teller, energy dispersive X‐ray, Zeta potential and particle size distribution values and AFM (Atomic force microscopy) techniques. Specially, our desired nanomaterial which has FA moieties on the surface of Tb@KCC‐1‐NH2‐FA where interact with folate receptor (FR) which there is on the surface of the various cancer cells. For this purpose, fluorescence microscopy images were used to prove the uptake of FA based nanomaterial with FR‐positive MDA breast cancer and HT 29 colon cancer cells. Also HEK 293 normal cells as FR‐negative cells verified the specificity of our desired nanomaterial toward the FR‐positive cells. The cytotoxicity survey of Tb@KCC‐1‐NH₂‐FA was examined by MTT assays against MDA breast cancer, HT 29 colon cancer and HEK 293 Normal cell lines which confirmed their biocompatible nature with any significant cytotoxic effects even for concentration higher than 900 μg/mL which could be used as a non‐toxic catalyst or carrier in biological ambient. Hence, Tb@KCC‐1‐NH₂‐FA were synthesized using green and hydrothermal method; the process was simple with good productivity and desired nanocomposite was non‐toxic.     

Effects of trehalose and sorbitol on the activity and structure of Pseudomonas cepacia lipase: spectroscopic insight

The stability of enzymes with no reduction in their catalytic activity still remains a critical issue in industrial applications. Naturally occurring osmolytes are commonly used as protein stabilizers. In this study we have investigated the effects of sorbitol and trehalose on the structural stability and activity of Pseudomonas cepacia lipase (PCL), using UV-visible, circular dichroism (CD) and fluorescence spectroscopy. Surface plasmon resonance (SPR) technique was used to trace changes in the refractive index and dielectric constant of the environment. The results revealed that catalytic activity and intrinsic fluorescence intensity of PCL increased in the presence of both osmolytes. Far-UV CD spectra indicated that the protein has undergone some conformational changes upon interacting with these osmolytes. Increasing the concentration of sorbitol led to changes in the refractive index and consequently the dielectric constant of environment; whereas in the case of trehalose, such changes were not significant. Unfavorable interactions of trehalose with protein surface induced higher preferential exclusion from the enzyme-water interface than that of sorbitol. Results of this report could give further insights about the stabilization mechanism of osmolytes.