Genetic diversity, population structure and marker-trait associations in Indian kale (Brassica oleracea L. gp. acephala) using cross-species microsatellite markers.

Kale is known for its exceptional nourishing and functional benefits to human body. However, it is an understudied species from genomic as well as agronomic aspects. It is important to characterize niche kale germplasms around the world to systematically conserve and utilize its genetic variability, especially for commercial traits in the interest of growers, consumers and industry. With this view, genomic and phenotypic characterizations of 62 Kashmiri kale accessions including popular landraces were done to estimate and partition genetic diversity, understand trait relationships, develop population structure and divulge marker-trait associations of economic significance. Sixty-six cross species microsatellite (SSR) markers within Brassica genus amplified 269 alleles in the germplasm. Their polymorphic information content (PIC) ranged from 0.00078 to 0.953 with an average of 0.407. The population structure analysis and neighbour joining tree clustering categorized the germplasm into three sub-populations. AMOVA revealed more within-population variance (67.73 %) than among-populations (32.27 %) variance. The principal component analysis (PCA) involving 24 agronomical traits revealed seven PCs (PC1 to PC7) having Eigen values more than 1, which explained a cumulative variation of 69.21 %. Association mapping with respect to these 24 agronomical traits using mixed linear model and general linear model revealed six overlapping significant marker-trait relationships with five being significant at probability value of 0.001/0.0001. The highly significant associations of two SSRs with economically important traits (siliqua length and seed weight) significantly correlated/related with leaf yield and seed yield were revealed for their possible utilization in marker assisted breeding for higher leaf and seed yields.

Derivation of mathematical closed form expressions for certain irregular topological indices of 2D nanotubes.

A numeric quantity that characterizes the whole structure of a network is called a topological index. In the studies of QSAR and QSPR, the topological indices are utilized to predict the physical features related to the bioactivities and chemical reactivity in certain networks. Materials for 2D nanotubes have extraordinary chemical, mechanical, and physical capabilities. They are extremely thin nanomaterials with excellent chemical functionality and anisotropy. Since, 2D materials have the largest surface area and are the thinnest of all known materials, they are ideal for all applications that call for intense surface interactions on a small scale. In this paper, we derived closed formulae for some important neighborhood based irregular topological indices of the 2D nanotubes. Based on the obtained numerical values, a comparative analysis of these computed indices is also performed.

Microplastic pollution in the surface water and sediments from Kallar Kahar wetland, Pakistan: occurrence, distribution, and characterization by ATR-FTIR.

This study reports the distribution of microplastics in surface water and sediments collected from Kallar Kahar wetland, Punjab, Pakistan, which is a game reserve and hosts migratory birds during winter season. Microplastics were extracted using density separation and wet oxidation method. The microplastics identification was done under a stereo-microscope, and their polymer compositions were characterized using an attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. The average abundance of microplastics in water and sediment samples was 88 ± 14.5 items/L and 5720 ± 2580 items/kg, respectively. The dominant shape groups of microplastics in water were fiber (58.7%), irregular fragments (32.4%), and beads (8.7%) with dominant colors as transparent > black > yellow ≈ white > red > green > pink > blue. Similar distribution in sediments was found, i.e., fiber (61.2%), irregular fragments (28.4%), and beads (10.3%) with dominant colors as transparent > pink > white > red ≈ black > blue > brown > green ≈ yellow. The ATR-FTIR spectra of visible microplastics were identified to be polypropylene (PP), high density polyethylene (HDPE), ethylene vinyl acetate (EVA), low density polyethylene (LDPE), nitrile, polymethyl methacrylate (PMMA), latex, and polyethylene terephthalate (PETE). In the study area, recreational activities, improper waste disposal, and runoff from catchment areas are the main reasons for the contamination of microplastics in the Lake. The pollution load can be minimized by taking measures such as creating awareness, promotion of ecotourism, and reducing plastic use.

Economic growth, social inclusion, and environmental protection: assessing the existence of green growth in Pakistan.

The Fifth Ministerial Conference on Environment and Development in Asia and the Pacific (MCED-5) provided a regional implementation plan to pursue economic development in Asia-Pacific. Achieving environmentally sustainable economic growth or green growth is considered necessary by the ministerial declaration of the conference. The ministerial declaration defines green growth as an approach to sustaining economic growth and employment creation, a prerequisite for effective poverty reduction while coping with natural resource constraints and climate change. Based on the importance of green growth, the study seeks to investigate the progress towards sustainable economic development in Pakistan from 1990 to 2019. The study employs structural equation modeling (SEM) to determine the direct and indirect effects of the variables of the green growth model adopted in the MCED-5. The results of the study indicate that an increase in the net national income of the country leads to increased natural resource depletion. The declining stock of natural capital points towards the difficulty in fulfilling biocapacity sustainability in Pakistan while achieving social progress and declining carbon intensity in the quest for sustainable development. Based on the analysis, it can be claimed that the negative impact of increasing inclusive wealth on natural capital makes Pakistan in environmental terms a weakly sustainable nation. Thus, the conclusion is that Pakistan is following a path of weak sustainability. As a result, there is a need to shift the country's sustainable economic development from weak sustainability to strong sustainability if the increasing natural resource depletion is to be restrained.

A triple test cross analysis to detect epistatic gene effects in cabbage (Brassica oleracea var. capitata L.): An updated methodology.

The cross-pollinated crop Brassica oleracea var. capitata L. shows good heterotic heterosis at high output; better standing of the plants; early maturity; larger and more homogeneous heads; consistency of head compactness; and disease-tolerance in F1 hybrids. There is very limited information documented on the epistasis of essential cabbage characters. We expand the research in this study to include an upgraded test to cross-design for enrolling and estimating epistasis and other genetic variance components controlling head yield and component traits in cabbage. The data was obtained from 45 families produced by crossing 15 lines with three testers; SC 2008-09, E-1-3-1&2, and their single cross F1, was subjected to triple test cross analysis. The current study results confirmed "j + 1" form of epistasis which is a major component for all traits. The plant spread, non-wrapper leaves, nethead/grossweight, polar/equatorial diameter, marketable head yield per plot, iron content and dry matter lugged both "j + 1" and 'i' type with the predominance of the 'i' type of interaction. Except for head shape index, equatorial diameter, head compactness was more noticeable when observed in dominance component. The degree of dominance is in the partial range, but both the head shape index/compactness and equatorial diameter showed over dominance. For maximum part, superiority was shown in both the directions. Appropriate breeding procedures are proposed to exploit the different forms of gene effects discovered for genetic improvement of head yield and quality traits.

Palyno-morphological characterization of selected allergenic taxa of family Poaceae from Islamabad-Pakistan using microscopic techniques.

Statistics around the world show an increased risk of high allergies associated with plants. The role of allergenic pollen is now well known. The most serious allergenic diseases are asthma and rhinitis, which affect 5-30% of the population of industrialized countries. This work focuses on the exploration of allergenic grass species of Islamabad, Pakistan and use microscopic techniques to correctly identify them. This study contains 11 species of grasses which showed variations in pollen morphological features. The dominant pollen shape was prolate, prolate-spheroidal and sub oblate. The largest polar diameter was calculated for Echinochloa crus-galli (56.00 µm) while the largest equatorial diameter was found in Cenchrus ciliaris (45.00 µm). Exine ornamentation varied from Gemmate-verrucate, Scabrate-reugulate, Verrucate-reticulate, Gemmate-Scabrate, Fine-Gemmate, Gemmate-rugulate, Gemmate-reticulate, Gemmate-scabrate, Scabrate-verrucate, and Scabrate. Maximum exine thickness was found in Eleusine indica (4.90 µm) while in C. ciliaris (1.50 µm) the minimum thickness of exine was measured. In E. crus-galli (1.54 µm), the largest P to E value was measured and the smallest value was noticed in Setaria verticillata (0.86 µm). The study was performed with an aim to identify the allergenic plant species in Islamabad-Pakistan using pollen morphological studies. This study provides significant information, to arrange the pollen types within their defined classes and to discuss various pollen features for developing an allergenic pollen atlas.

Implication of scanning electron microscopy in the seed morphology with special reference to three subfamilies of Fabaceae.

The current research work was aimed to determine the morphological variation in the seeds of subfamilies Caesalpinioideae, Mimosoideae, and Papilionoideae using scanning electron microscope to investigate the different seed features including; Seed length, width, weight, hilum shape, position, fracture line pattern, pleurogram shape, texture crudeness, ornamentation wall, and surface ornamentation. A total of 12 species were collected. The study reveals that the seed types alters from psilate to rugose, pleurogram shape from lunar to U shape, hilum shape from elliptical to depressed, hilum position from terminal to subterminal, fracture line pattern from reticulate irregular ridges to psilate with regular ridges, texture crudeness from fine to coarse, surface ornamentation from reticulate to psilate, and ornamentation wall also varies from thin to thick. In all the studied species, it was found that seed surface was brown in color except in Senna alata, in which seed surface was black. Regarding the seed size, it was found that the smallest seed was observed for Sesbania sesban, while the largest seed was observed for Pongamia pinnata. The present research article indicates the taxonomic significance of seed morphology in the subfamilies Caesalpinioideae, Mimosaceae, and Papilionoideae.

Synthesis, characterization and docking studies of amide ligands as anti-leishmanial agents.

Aim of this study was to synthesize new inhibitors on the basis of active site of aspartic protease enzyme and to evaluate their intended biological activity. A3D model of an enzyme was generated via homology modeling and series of novel amide ligands were synthesized by using a short high yield process, subsequently, analyzed in-silico and in-vitro anti-leishmanial activities. Characterization and identification was accomplished via NMR (H1& C13), infrared and mass spectroscopic techniques. Among all compound (4) was found to show significant activity (IC50 58±0.01) against Leishmania major (L. major) species. Furthermore, docking studies confirmed the inhibition of a targeted enzyme that supported the interaction of potent compound (4) with key residues (aspartic protease) via hydrogen bonds. Present study conferred about novel compound (4) as a promising compound to antagonize L. major activities in future.

Investigating Schizocarp morphology as a taxonomic tool in study of Apiaceae family by utilizing LM and SEM techniques.

In present study, the schizocarp morphology of 14 species belonging to Apiaceae family has been investigated. Light microscopy (LM) and scanning electron microscopy (SEM) have been utilized to highlight qualitative and quantitative features of studied species. Variations have been observed in macro- and micro-morphological features such as color, shape, symmetry, length, width, apex, epicuticular projections, surface patterns, anticlinal, and periclinal wall patterns. Schizocarp shapes observed were oval, round, triangular, linear, elliptic, and globose. Fruit was either homomorphic or heteromorphic. Crystalloids, stellate hair, multicellular spines, and platelets were mostly observed epicuticular projections. Surface patterns on the fruit surface were striate, rugulate-striate, reticulate, and striato-knotted. Both macro- and micro-morphological characters can serve as an important tool in classifying Apiaceae family at various taxonomic ranks. Substantial variations observed can assist as useful constraints at various taxonomic levels as they provide reliable and constant details. Disparities observed in schizocarp features can pave a path for Apiaceae family classification based on phylogenetic and molecular studies.

Macromorphological and micromorphological seed features of selected taxa of Caesalpiniaceae using light and scanning electron microscopy.

Seed micromorphological and macromorphological characteristics of 12 taxa of Caesalpiniaceae from Pakistan have been studied, using light and scanning electron microscopy, to investigate the importance of seed coat features as a taxonomic tool. Great variations have been observed in color, shape, dimension, and seed surface pattern among the different genera of the family. A taxonomic key was prepared for the studied taxa. Six types of seed shapes were observed; circular, elliptical, irregular, oblong, oval, and ovoid. All examined seeds were hard except fragile seeds of Haematoxylom campechianum. Eight types of surface ornamentation have been noticed that include levigate, lugose, papillate, reticulate, reticulate irregular, reticulate regular, rhombus, and rogues. Majority of the taxa has been observed with thick ornamentation wall but thin ornamentation wall has also been recorded in few species. Fracture line of the various patterns was present in all taxa except genus Bauhinia. Three types of texture crudeness; coarse, medium, and fine have been recorded. Both micromorphological and macromorphological characters of seed are very fruitful in identification and classification of Caesalpiniaceae.

Preimplantation Factor (PIF) Promotes HLA-G, -E, -F, -C Expression in JEG-3 Choriocarcinoma Cells and Endogenous Progesterone Activity.

BACKGROUND/AIMS: Pregnancy success requires mandatory maternal tolerance of the semi/ allogeneic embryo involving embryo-derived signals. Expression levels of PreImplantation Factor (PIF), a novel peptide secreted by viable embryos, correlate with embryo development, and its early detection in circulation correlates with a favourable pregnancy outcome. PIF enhances endometrial receptivity to promote embryo implantation. Via the p53 pathway, it increases trophoblast invasion, improving cell survival / immune privilege. PIF also reduces spontaneous and LPS-induced foetal death in immune naïve murine model. We examined PIF effect on gene expression of human leukocyte antigen (HLA-G, -E -F and -C) and the influence of PIF on local progesterone activity in JEG-3 choriocarcinoma cells. METHODS: PIF and progesterone (P4) effects on JEG-3 cells surface and intracellular HLA molecules was tested using monoclonal antibodies, flow cytometry, and Western blotting. PIF and IL17 effects on P4 and cytokines secretion was determined by ELISA. PIF and P4 effects on JEG-3 cells proteome was examined using 2D gel staining followed by spot analysis, mass spectrometry and bioinformatic analysis. RESULTS: In cytotrophoblastic JEG-3 cells PIF increased intracellular expression of HLA-G, HLA-F, HLA-E and HLA-C and surface expression of HLA-G, HLA-E and HLA-C in dose and time dependent manner. In case of HLA-E, -F results were confirmed also by Western blot. Proteome analysis confirmed an increase in HLA-G, pro-tolerance FOXP3+ regulatory T cells (Tregs), coagulation factors and complement regulator. In contrast, PIF reduced PRDX2 and HSP70s to negate oxidative stress and protein misfolding. PIF enhanced local progesterone activity, increasing steroid secretion and the receptor protein. It also promoted the secretion of the Th1/Th2 cytokines (IL-10, IL-1ß, IL-8, GM-CSF and TGF-ß1), resulting in improved maternal signalling. CONCLUSION: PIF can generate a pro-tolerance milieu by enhancing the expression of HLA molecules and by amplifying endogenous progesterone activity. A Fast-Track clinical trial for autoimmune disease has been satisfactorily completed. The acquired data warrants PIF use for the treatment of early pregnancy disorders.

Activated Cdc42-associated kinase 1 (ACK1) binds the sterile α motif (SAM) domain of the adaptor SLP-76 and phosphorylates proximal tyrosines.

The adaptor protein Src homology 2 domain-containing leukocyte phosphoprotein of 76 kDa (SLP-76) plays a crucial role in T cell activation by linking antigen receptor (T cell receptor, TCR) signals to downstream pathways. At its N terminus, SLP-76 has three key tyrosines (Tyr-113, Tyr-128, and Tyr-145, "3Y") as well as a sterile α motif (SAM) domain whose function is unclear. We showed previously that the SAM domain has two binding regions that mediate dimer and oligomer formation. In this study, we have identified SAM domain-carrying non-receptor tyrosine kinase, activated Cdc42-associated tyrosine kinase 1 (ACK1; also known as Tnk2, tyrosine kinase non-receptor 2) as a novel binding partner of SLP-76. Co-precipitation, laser-scanning confocal microscopy, and in situ proximity analysis confirmed the binding of ACK1 to SLP-76. Further, the interaction was induced in response to the anti-TCR ligation and abrogated by the deletion of SLP-76 SAM domain (ΔSAM) or mutation of Tyr-113, Tyr-128, and Tyr-145 to phenylalanine (3Y3F). ACK1 induced phosphorylation of the SLP-76 N-terminal tyrosines (3Y) dependent on the SAM domain. Further, ACK1 promoted calcium flux and NFAT-AP1 promoter activity and decreased the motility of murine CD4+ primary T cells on ICAM-1-coated plates, an event reversed by a small molecule inhibitor of ACK1 (AIM-100). These findings identify ACK1 as a novel SLP-76-associated protein-tyrosine kinase that modulates early activation events in T cells.

Effect of Enzymatic pre-treatment of microalgae extracts on their anti-tumor activity.

BACKGROUND: There is an increasing need to find natural bioactive compounds for pharmaceutical applications, because they have less harmful side effects compared to their chemical alternatives. Microalgae (MA) have been identified as a promising source for these bioactive compounds, and this work aimed to evaluate the anti-proliferative effects of semi-purified protein extracted from MA against several tumor cell lines. METHODS: Tested samples comprised MA cell extracts treated with cellulase and lysozyme, prior to extraction. The effect of dialysis, required to remove unnecessary small molecules, was also tested. The anti-cancer efficacies of the dialyzed and undialyzed extracts were determined by measuring cell viability after treating four human cancer cell lines, specifically A549 (human lung carcinoma), MCF-7 (human breast adenocarcinoma), MDA MB-435 (human melanoma), and LNCap (human prostate cancer cells derived from a metastatic site in the lymph node). This was compared to the effects of the agents on the human BPH-1 cell line (benign human prostate epithelial cells). The t-test was used to statistically analyze the results and determine the significance. RESULTS: Against LNCap and A549 cells, the performance of cellulase-treated extracts was better (with p-values < 0.05, as compared to the control) than that of lysozyme-treated preparations (with p-values mainly > 0.05, as compared to the control); however, they had similar effects against the other two tumor cell lines (with p-values mainly < 0.05, as compared to the control). Moreover, based on their effect on BPH-1 cells, extracts from lysozyme-treated MA cells were determined to be safer against the benign prostate hyperplasia cells, BPH-1 (with p-values mainly > 0.05, as compared to the control). After dialysis, the performance of MA extracts from lysozyme-treated cells was enhanced significantly (with p-values dropping to < 0.05, as compared to the control). CONCLUSIONS: The results of this work provide important information and could provide the foundation for further research to incorporate MA constituents into pharmaceutical anti-cancer therapeutic formulations.

Sertoli cells have a functional NALP3 inflammasome that can modulate autophagy and cytokine production.

Sertoli cells, can function as non-professional tolerogenic antigen-presenting cells, and sustain the blood-testis barrier formed by their tight junctions. The NOD-like receptor family members and the NALP3 inflammasome play a key role in pro-inflammatory innate immunity signalling pathways. Limited data exist on NOD1 and NOD2 expression in human and mouse Sertoli cells. Currently, there is no data on inflammasome expression or function in Sertoli cells. We found that in primary pre-pubertal Sertoli cells and in adult Sertoli line, TLR4\NOD1 and NOD2 crosstalk converged in NFκB activation and elicited a NALP3 activation, leading to de novo synthesis and inflammasome priming. This led to caspase-1 activation and IL-1ß secretion. We demonstrated this process was controlled by mechanisms linked to autophagy. NOD1 promoted pro-IL-1ß restriction and autophagosome maturation arrest, while NOD2 promoted caspase-1 activation, IL-1ß secretion and autophagy maturation. NALP3 modulated NOD1 and pro-IL-1ß expression, while NOD2 inversely promoted IL-1ß. This study is proof of concept that Sertoli cells, upon specific stimulation, could participate in male infertility pathogenesis via inflammatory cytokine induction.

Quantified colocalization reveals heterotypic histocompatibility class I antigen associations on trophoblast cell membranes: relevance for human pregnancy.

Human placental syncytiotrophoblasts lack expression of most types of human leukocyte antigen (HLA) class I and class II molecules; this is thought to contribute to a successful pregnancy. However, the HLA class Ib antigens HLA-G, -E, and -F and the HLA class Ia antigen HLA-C are selectively expressed on extravillous trophoblast cells, and they are thought to play a major role in controlling feto-maternal tolerance. We have hypothesized that selective expression, coupled with the preferential physical association of pairs of HLA molecules, contribute to the function of HLA at the feto-maternal interface and the maternal recognition of the fetus. We have developed a unique analytical model that allows detection and quantification of the heterotypic physical associations of HLA class I molecules expressed on the membrane of human trophoblast choriocarcinoma cells, ACH-3P and JEG-3. Automated image analysis was used to estimate the degree of overlap of HLA molecules labeled with different fluorochromes. This approach yields an accurate measurement of the degree of colocalization. In both JEG-3 and ACH-3P cells, HLA-C, -E, and -G were detected on the cell membrane, while the expression of HLA-F was restricted to the cytoplasm. Progesterone treatment alone induced a significant increase in the expression level of the HLA-G/HLA-E association, suggesting that this heterotypic association is modulated by this hormone. Our data shows that the cell-surface HLA class I molecules HLA-G, -E, and -C colocalize with each other and have the potential to form preferential heterotypic associations.

A novel method for quantified, superresolved, three-dimensional colocalisation of isotropic, fluorescent particles.

Colocalisation, the overlap of subcellular structures labelled with different colours, is a key step to characterise cellular phenotypes. We have developed a novel bioimage informatics approach for quantifying colocalisation of round, blob-like structures in two-colour, highly resolved, three-dimensional fluorescence microscopy datasets. First, the algorithm identifies isotropic fluorescent particles, of relative brightness compared to their immediate neighbourhood, in three dimensions and for each colour. The centroids of these spots are then determined, and each object in one location of a colour image is checked for a corresponding object in the other colour image. Three-dimensional distance maps between the centroids of differently coloured spots then display where and how closely they colocalise, while histograms allow to analyse all colocalisation distances. We use the method to reveal sparse colocalisation of different human leukocyte antigen receptors in choriocarcinoma cells. It can also be applied to other isotropic subcellular structures such as vesicles, aggresomes and chloroplasts. The simple, robust and fast approach yields superresolved, object-based colocalisation maps and provides a first indication of protein-protein interactions of fluorescent, isotropic particles.

Stem cells in the reproductive system.

This review article summarizes current knowledge on regulation, functions, and capacities of stem cells in the female and male reproductive tract. Major locations in which pluripotent cells reside and from where they can be isolated are the ovaries, the endometrium, the decidua, and the testis. They include oocytes, embryonic stem cells, trophoblast stem cells, and spermatogonial stem cells, but also several side populations, which can be obtained after certain isolation and culture procedures. The potential of pluripotent cells in the reproductive tract to differentiate is manifold, but heterogenous, depending upon their respective origin. As stem cells have a potential for future application in transplantation and regenerative medicine, this article also reviews the literature on major histocompatibility complex expression on stem cells of the reproductive tract, because of its immunogenic effects, but also because of its potential expression of HLA-G, a potent immunomodulator mainly associated with trophoblast cells.

Key cellular components and interactive histocompatibility molecules regulating tolerance to the fetal allograft.

Implantation is a major landmark in life. It involves the correct apposition of the embryo in the maternal endometrium. The cellular environment influences placenta development, and direct contact of the fetus with maternal tissues is achieved through decidual cells. At the decidua, and at systemic level, the correct balance of cells potentially acting as antigen-presenting cells and histocompatibility products play a pivotal role in achieving feto-maternal tolerance. Here, we review some of the current issues associated with the interplay between cells and molecules needed for pregnancy development.

Effects of simvastatin and alpha-tocopherol on disturbed nerve conduction in obese Sprague Dawley rats.

BACKGROUND: The incidence of obesity is increasing worldwide. The neuropathy associated with obesity, that is evident from disturbed nerve conduction, is one of the complications for which a number of treatment options are being considered. In this study, Simvastatin, a hydroxyl methyl glutaryl coenzyme A reductase inhibitor and alpha-tocopherol, a dietary antioxidant are compared for their effects on sciatic nerve conduction velocity. OBJECTIVES: To compare the effects of Simvastatin and alpha-tocopherol on sciatic nerve conduction velocity in obese rats. METHODS: The study was a Randomised control trial conducted from December 2008 to November 2009. One hundred and twenty adult male Sprague Dawley rats were divided into four groups with 30 rats in each group. One group of rats was taken as control with normal diet while other three groups were given high fat diet (HFD) for the whole study period. Along with the high fat diet, group III and group IV were given Simvastatin and alpha-tocopherol supplemented diet respectively. At the end of study, conduction velocity of sciatic nerve was determined with the help of PowerLab data acquisition system. RESULTS: The three groups with HFD showed more than 25% increase in weight at the end of study compared to control group. The control group with high fat diet (Group II) showed decreased sciatic nerve conduction velocity when compared with control (Group I). Both the groups that were given Simvastatin and alpha-tocopherol each showed improvement in sciatic nerve conduction velocity (p<0.001) after four weeks when compared with the group that was given HFD without any supplementation. However with alpha-tocopherol, the nerve conduction velocity was improved more significantly. CONCLUSIONS: Simvastatin and alpha-tocopherol both are effective for improving sciatic nerve conduction velocity in HFD induced obesity.