Microengineered systems with iPSC-derived cardiac and hepatic cells to evaluate drug adverse effects

Hepatic and cardiac drug adverse effects are among the leading causes of attrition in drug development programs, in part due to predictive failures of current animal or in vitro models. Hepatocytes and cardiomyocytes differentiated from human induced pluripotent stem cells (iPSCs) hold promise for predicting clinical drug effects, given their human-specific properties and their ability to harbor genetically determined characteristics that underlie inter-individual variations in drug response. Currently, the fetal-like properties and heterogeneity of hepatocytes and cardiomyocytes differentiated from iPSCs make them physiologically different from their counterparts isolated from primary tissues and limit their use for predicting clinical drug effects. To address this hurdle, there have been ongoing advances in differentiation and maturation protocols to improve the quality and use of iPSC-differentiated lineages. Among these are in vitro hepatic and cardiac cellular microsystems that can further enhance the physiology of cultured cells, can be used to better predict drug adverse effects, and investigate drug metabolism, pharmacokinetics, and pharmacodynamics to facilitate successful drug development. In this article, we discuss how cellular microsystems can establish microenvironments for these applications and propose how they could be used for potentially controlling the differentiation of hepatocytes or cardiomyocytes. The physiological relevance of cells is enhanced in cellular microsystems by simulating properties of tissue microenvironments, such as structural dimensionality, media flow, microfluidic control of media composition, and co-cultures with interacting cell types. Recent studies demonstrated that these properties also affect iPSC differentiations and we further elaborate on how they could control differentiation efficiency in microengineered devices. In summary, we describe recent advances in the field of cellular microsystems that can control the differentiation and maturation of hepatocytes and cardiomyocytes for drug evaluation. We also propose how future research with iPSCs within engineered microenvironments could enable their differentiation for scalable evaluations of drug effects.     

Dynamics of solute/matric stress interactions with climate change abiotic factors on growth,gene expression and ochratoxin A production by Penicillium verrucosum on a wheat-based matrix

Penicillium verrucosum contaminates temperate cereals with ochratoxin A (OTA) during harvesting and storage. We examined the effect of temperature (25 vs 30 ^oC), CO₂ (400 vs 1000 ppm) and matric/solute stress (-2.8 vs -7.0 MPa) on (i) growth, (ii) key OTA biosynthetic genes and (iii) OTA production on a milled wheat substrate. Growth was generally faster under matric than solute stress at 25 ^oC, regardless of CO₂ concentrations. At 30 ^oC, growth of P. verrucosum was significantly reduced under solute stress in both CO₂ treatments, with no growth observed at -2.8 MPa (=0.98 water activity, a_w) and 1000 ppm CO₂. Overall, growth patterns under solute stress was slower in elevated CO₂ than under matric stress when compared with existing conditions. The otapksPV gene expression was increased under elevated CO₂ levels in matric stress treatments. There was fewer effects on the otanrpsPV biosynthetic gene. This pattern was paralleled with the production of OTA under these conditions. This suggest that P. verrucosum is able to actively grow and survive in both soil and on crop debris under three way interacting climate-related abiotic factors. This resilience suggests that they would still be able to pose an OTA contamination risk in temperate cereals post-harvest.     

Perspectives of pluripotent stem cells in livestock

The recent progress in derivation of pluripotent stem cells(PSCs)from farm animals opens new approaches not only for reproduction,genetic engineering,treatment and conservation of these species,but also for screening novel drugs for their efficacy and toxicity,and modelling of human diseases.Initial attempts to derive PSCs from the inner cell mass of blastocyst stages in farm animals were largely unsuccessful as either the cells survived for only a few passages,or lost their cellular potency;indicating that the protocols which allowed the derivation of murine or human embryonic stem(ES)cells were not sufficient to support the maintenance of ES cells from farm animals.This scenario changed by the innovation of induced pluripotency and by the development of the 3 inhibitor culture conditions to support na?ve pluripotency in ES cells from livestock species.However,the long-term culture of livestock PSCs while maintaining the full pluripotency is still challenging,and requires further refinements.Here,we review the current achievements in the derivation of PSCs from farm animals,and discuss the potential application areas.     

Purification, Spectroscopic Characterization and o-Diphenoloxidase Activity of Hemocyanin from a Freshwater Gastropod: Pila globosa

Hemocyanins are multi-subunit oxygen carrier proteins, found in select species of arthropoda and mollusca. Here, we have purified native hemocyanin from Pila globosa, a freshwater gastropod, verified using mass spectrometry and determined its molecular weight, secondary structure and the spectral properties, using Ultraviolet/visible, Fourier transform infra-red and Circular dichroism spectroscopy. Our results reveal the oligomeric and glycosylated nature of the protein, comprising of 400 kDa subunits, organized predominantly into a thermo-stable, alpha-helical conformation. Further, biochemical assays confirm catecholoxidase-like activity in hemocyanin, which has been used to develop a first-generation optical sensor, for the detection of phenols.     

Modifying bio-catalytic properties of enzymes for efficient biocatalysis: a review from immobilization strategies viewpoint

Enzyme-based catalysis has become one of the most important disciplines in organic synthesis and plays a noteworthy role in the establishment of many chemical industries, e.g. fine chemicals, food or energy, textiles, agricultural, cosmeceutical, medicinal and pharmaceutical industries. However, pristine enzymes fail to demonstrate requisite functionalities for an industrial setting where extremely specific and stable catalysts are required. Immobilization enhances the catalytic stability and activity of enzymes and trims the overall cost burden of the enzyme. Therefore, it widely endeavours for proficient, sustainable, and environmentally responsive catalytic processes. Amongst several immobilization strategies, e.g. (1) supports-assisted, i.e. physical or covalent coupling and (2) supports-free techniques, i.e. cross-linked enzyme crystals (CLECs) or aggregates are the most promising ones and widely pursued for enzyme immobilization purposes. This perspective review focuses on up-to-date developments in the area of enzyme immobilization and presents their potentialities to upgrade and/or modify enzyme properties. Both types of immobilization strategies, i.e. supports-assisted and supports-free techniques are discussed with particular reference to CLECs or aggregates and protein-coated microcrystals. Also, several useful traits achieved after immobilization are also discussed in the second half of the review.     

High serum free fatty acids and low leptin levels: Plausible metabolic indicators of negative energy balance in early lactating Murrah buffaloes

Lactation is a highly demanding event in mammals, including buffaloes. It modulates the partitioning of nutrients, energy utilization, and food intake of the mother to meet her own and infant's energy needs. Failure to satisfy these energy needs leads to Negative Energy Balance (NEB). Currently, the only available indirect NEB indicator is Body Condition Score (BCS). However, direct dependency of the BCS on the peak depletion of body fat causes its inefficient use in a dairy farm. Thus, to establish objective NEB indicators in buffaloes, the serum levels of biochemical (serum β-hydroxybutyrate [BHBA] and free fatty acids [FFAs]), and endocrine (Growth Hormone [GH], insulin-like growth factor1 [IGF1], Insulin, and leptin) parameters were estimated in buffaloes. Our results revealed that serum FFA levels were significantly (p < 0.05) higher in high milk yielders (HMY) than low milk yielders (LMY) and heifers (H) during the 3rd and the 4th weeks of postpartum. The serum FFA levels were also significantly (p < 0.001) higher in the postpartum buffaloes with BCS < 3 in the field conditions. Further, serum leptin levels were significantly (p < 0.05) lower in HMY than LMY during the 3rd week of postpartum. However, the BHBA, GH, IGF1, and insulin levels were not significantly different between lactating buffaloes and H. These observations indicated that the NEB condition is probably restricted to the first month of early lactation in buffaloes. In conclusion, the simultaneous higher FFA and lower leptin levels could act as direct plausible metabolic indicators of NEB in buffaloes.     

Fishing cat Prionailurus viverrinus distribution and habitat suitability in Nepal

The fishing cat Prionailurus viverrinus is a wetland specialist species endemic to South and Southeast Asia. Nepal represents the northern limit of its biogeographic range, but comprehensive information on fishing cat distribution in Nepal is lacking. To assess their distribution, we compiled fishing cat occurrence records (n = 154) from Nepal, available in published literature and unpublished data (2009–2020). Bioclimatic and environmental variables associated with their occurrence were used to predict the fishing cat habitat suitability using MaxEnt modeling. Fishing cat habitat suitability was associated with elevation (152–302 m), precipitation of the warmest quarter, i.e., April–June (668–1014 mm), precipitation of the driest month (4–7 mm), and land cover (forest/grassland and wetland). The model predicted an area of 4.4% (6679 km²) of Nepal as potential habitat for the fishing cat. About two‐thirds of the predicted potentially suitable habitat lies outside protected areas; however, a large part of the highly suitable habitat (67%) falls within protected areas. The predicted habitat suitability map serves as a reference for future investigation into fishing cat distribution as well as formulating and implementing effective conservation programs in Nepal. Fishing cat conservation initiatives should include habitats inside and outside the protected areas to ensure long‐term survival. We recommend conservation of wetland sites, surveys of fishing cats in the identified potential habitats, and studying their genetic connectivity and population status.     

Habitat occupancy of sloth bear Melursus ursinus in Chitwan National Park,Nepal

Mammals have experienced a massive decline in their populations and geographic ranges worldwide. The sloth bear, Melursus ursinus (Shaw, 1791), is one of many species facing conservation threats. Despite being endangered in Nepal, decades of inattention to the situation have hindered their conservation and management. We assessed the distribution and patterns of habitat use by sloth bears in Chitwan National Park (CNP), Nepal. We conducted sign surveys from March to June, 2020, in 4 × 4 km grids (n = 45). We collected detection/non‐detection data along a 4‐km trail that was divided into 20 continuous segments of 200 m each. We obtained environmental, ecological, and anthropogenic covariates to understand determinants of sloth bear habitat occupancy. The data were analyzed using the single‐species single‐season occupancy method, with a spatially correlated detection. Using repeated observations, these models accounted for the imperfect detectability of the species to provide robust estimates of habitat occupancy. The model‐averaged occupancy estimate for the sloth bear was 69% and the detection probability was 0.25. The probability of habitat occupancy by sloth bears increased with the presence of termites and fruits and in rugged, dry, open, undisturbed habitats. Our results indicate that the sloth bear is elusive, functionally unique, and widespread in CNP. Future conservation interventions and action plans aimed at sloth bear management must adequately consider their habitat requirements.     

Crystalline and water soluble CR(4+) and CR(5+:) model compounds for chromium toxicity studies

We discuss two complexes of Cr(4+) for their possible utility as models for Cr toxicity studies. They are Cr(dien)(O2)2(.)H2O (dien = diethylenetriamine) and Cr(NH3)3(O2)2, which have been recently characterized by x-ray diffraction and magnetic techniques. We present their optical and infrared absorption spectra as quick aids in their identification procedure. We also summarize the general properties of some well-characterized Cr(5+) compounds. All of these compounds are water soluble with the Cr(5+) samples being fairly stable in basic solutions. They can all be prepared as pure crystals with a shelf life of over 2 years when stored in a refrigerator.