An AFM Approach Applied in a Study of α-Crystallin Membrane Association: New Insights into Lens Hardening and Presbyopia Development

The lens of the eye loses elasticity with age, while α-crystallin association with the lens membrane increases with age. It is unclear whether there is any correlation between α-crystallin association with the lens membrane and loss in lens elasticity. This research investigated α-crystallin membrane association using atomic force microscopy (AFM) for the first time to study topographical images and mechanical properties (breakthrough force and membrane area compressibility modulus (KA), as measures of elasticity) of the membrane. α-Crystallin extracted from the bovine lens cortex was incubated with a supported lipid membrane (SLM) prepared on a flat mica surface. The AFM images showed the time-dependent interaction of α-crystallin with the SLM. Force spectroscopy revealed the presence of breakthrough events in the force curves obtained in the membrane regions where no α-crystallin was associated, which suggests that the membrane’s elasticity was maintained. The force curves in the α-crystallin submerged region and the close vicinity of the α-crystallin associated region in the membrane showed no breakthrough event within the defined peak force threshold, indicating loss of membrane elasticity. Our results showed that the association of α-crystallin with the membrane deteriorates membrane elasticity, providing new insights into understanding the molecular basis of lens hardening and presbyopia

Effects of calcium channel blocker, nifedipine, on antidepressant activity of fluvoxamine, venlafaxine and tianeptine in mice

Background: Cardiovascular diseases are commonly associated with depression. Calcium channel blockers (CCBs) form commonly used group of drugs for the treatment of a number of cardiovascular diseases. Nifedipine, a CCB, has been shown to possess antidepressant activity and potentiate antidepressant activity of imipramine and sertraline, however, literature on its interaction with newer antidepressant drugs such as fluvoxamine, venlafaxine and tianeptine is limited. Hence, the present study was undertaken.Methods: The study was carried out in albino mice in two phases. In Phase I, antidepressant activity of nifedipine, fluvoxamine, venlafaxine and tianeptine were confirmed after their single dose administration using forced swim test (FST) and tail suspension test (TST) and their minimum antidepressant doses were determined. In Phase II, the effect of nifedipine on antidepressant activity of fluvoxamine, venlafaxine and tianeptine was studied by orally administering sub-antidepressant doses of these drugs for 28 days. FST and TST were carried out on 1st, 14th and 28th day of the study and change in immobility period was observed.Results: In Phase I, all the studied drugs exhibited dose dependent antidepressant activity in both FST and TST. Minimal antidepressant dose of nifedipine, fluvoxamine, venlafaxine and tianeptine was observed as 10, 25, 25 and 10 mg/kg respectively. In Phase II, combinations of sub-antidepressant dose of nifedipine (5 mg/kg) with sub-antidepressant doses of fluvoxamine (12.5 mg/kg), venlafaxine (12.5 mg/kg) and tianeptine (5 mg/kg) exhibited enhanced antidepressant activity when compared to the control group and individual drug groups after same duration of treatment.Conclusions: Nifedipine, fluvoxamine, venlafaxine and tianeptine possess antidepressant activity and nifedipine exhibits synergistic antidepressant activity with fluvoxamine, venlafaxine and tianeptine

Interaction of Alpha-Crystallin with Four Major Phospholipids of Eye Lens Membranes

It is well-studied that the significant factor in cataract formation is the association of α-crystallin, a major eye lens protein, with the fiber cell plasma membrane of the eye lens. The fiber cell plasma membrane of the eye lens consists of four major phospholipids (PLs), i.e., phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), and sphingomyelin (SM). Despite several attempts to study the interaction of α-crystallin with PLs of the eye lens membrane, the role of individual PL for the binding with α-crystallin is still unclear. We recently developed the electron paramagnetic resonance (EPR) spin-labeling method to study the binding of α-crystallin to the PC membrane (Mainali et al., 2020a). Here, we use the recently developed EPR method to explicitly measure the binding affinity (Ka) of α-crystallin to the individual (PE*, PS, and SM) and two-component mixtures (SM/PE, SM/PS, and SM/PC in 70:30 and 50:50 mol%) of PL membranes as well as the physical properties (mobility parameter and maximum splitting) of these membranes upon binding with α-crystallin. One of the key findings of this study was that the Ka of α-crystallin binding to individual PL membranes followed the trends: Ka(PC) \u3e Ka(SM) \u3e Ka(PS) \u3e Ka(PE*), indicating PE* inhibits binding the most whereas PC inhibits binding the least. Also, the Ka of α-crystallin binding to two-component mixtures of PL membranes followed the trends: Ka(SM/PE) \u3e Ka(SM/PS) \u3e Ka(SM/PC), indicating SM/PC inhibits binding the most whereas SM/PE inhibits binding the least. Except for the PE* membrane, for which there was no binding of α-crystallin, the mobility parameter for all other membranes decreased with an increase in α-crystallin concentration. It represents that the membranes become more immobilized near the headgroup regions of the PLs when more and more α-crystallin binds to them. The maximum splitting increased only for the SM and the SM/PE (70:30 mol%) membranes, with an increase in the binding of α-crystallin. It represents that the PL headgroup regions of these membranes become more ordered after binding of α-crystallin to these membranes. Our results showed that α-crystallin binds to PL membranes in a saturable manner. Also, our data suggest that the binding of α-crystallin to PL membranes likely occurs through hydrophobic interaction between α-crystallin and the hydrophobic fatty acid core of the membranes, and such interaction is modulated by the PL headgroup’s size and charge, hydrogen bonding between headgroups, and PL curvature. Thus, this study provides an in-depth understanding of α-crystallin interaction with the PL membranes made of individual and two-component mixtures of the four major PLs of the eye lens membranes

Water rights, conflict and policy: Proceedings of a workshop held in Kathmandu, Nepal, January 22-24, 1996

IrrigationIrrigation managementWater rightsWater lawConflictFarmers' associations

Soil Chemical Properties Under Conservation Agriculture and Cereal-Based Cropping System in Eastern Tarai of Nepal

Field experiments were conducted for four years (2014-2017) at five locations namely Salbani, Bhokraha, Simariya, Bhaluwa and Kaptanganj of Sunsari district to assess the changes in soil chemical properties under conservation agriculture (CA)-based practices in two cropping systems namely rice-kidney bean-maize at Salbani and rice-wheat at rest of the locations. In rice-wheat cropping system, there were four treatments: (1) conventional tillage (CT) for rice transplantation and subsequent wheat sowing, (2) conventional tillage rice transplantation followed by zero tillage (ZT) wheat, (3) unpuddled rice transplantation followed by zero tillage wheat, (4) zero tillage in both rice and wheat. Similarly, in rice-kidney bean-maize cropping system, there were four treatments; (1) conventional tillage for rice transplantation and sowing of both kidney bean and maize, (2) conventional tillage rice transplantation followed by zero tillage in both kidney bean and maize, (3) unpuddled rice transplantation followed by zero tillage in both kidney bean and maize, (4) zero tillage in all three crops. Soil samples were taken at initial and every year after rice harvest.The soil samples were analyzed for total nitrogen, available phosphorus, available potassium, pH and soil organic matter.Total nitrogen (N) showed a slightly decreasing trend in the first three years and showed a slight increase at the end of experiment under ZT in all locations. The total N under ZT changed from 0.12 to 0.13%, 0.05 to 0.06%, 0.10 to 0.12%, 0.11 to 0.08% and 0.09 to 0.13% in Salbani, Bhokraha, Simariya, Bhaluwa and Kaptanganj, respectively.  All locations showed the positive values of available potassium; Salbani  revealing considerable change of 64.3 to 78.5 mg/kg in CT while 68.4 to 73.3 mg/kg in ZT condition. The treatment where rice was transplanted in unpuddled condition and zero tilled to wheat, had a mean value of available phosphorus and potassium as 87.3 and 81.9 mg/kg respectively. Soil pH ranged from 4.8 to 7.1 in CT while it was 5.2 to 6.8 in ZT across the locations. The change in soil organic matter in CT of all locations except Salbani was narrower as compared to ZT

Interaction of β\u3csub\u3eL\u3c/sub\u3e- and γ-Crystallin with Phospholipid Membrane Using Atomic Force Microscopy

Highly concentrated lens proteins, mostly β- and γ-crystallin, are responsible for maintaining the structure and refractivity of the eye lens. However, with aging and cataract formation, β- and γ-crystallin are associated with the lens membrane or other lens proteins forming high-molecular-weight proteins, which further associate with the lens membrane, leading to light scattering and cataract development. The mechanism by which β- and γ-crystallin are associated with the lens membrane is unknown. This work aims to study the interaction of β- and γ-crystallin with the phospholipid membrane with and without cholesterol (Chol) with the overall goal of understanding the role of phospholipid and Chol in β- and γ-crystallin association with the membrane. Small unilamellar vesicles made of Chol/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (Chol/POPC) membranes with varying Chol content were prepared using the rapid solvent exchange method followed by probe tip sonication and then dispensed on freshly cleaved mica disk to prepare a supported lipid membrane. The βL- and γ-crystallin from the cortex of the bovine lens was used to investigate the time-dependent association of βL- and γ-crystallin with the membrane by obtaining the topographical images using atomic force microscopy. Our study showed that βL-crystallin formed semi-transmembrane defects, whereas γ-crystallin formed transmembrane defects on the phospholipid membrane. The size of semi-transmembrane defects increases significantly with incubation time when βL-crystallin interacts with the membrane. In contrast, no significant increase in transmembrane defect size was observed in the case of γ-crystallin. Our result shows that Chol inhibits the formation of membrane defects when βL- and γ-crystallin interact with the Chol/POPC membrane, where the degree of inhibition depends upon the amount of Chol content in the membrane. At a Chol/POPC mixing ratio of 0.3, membrane defects were observed when both βL- and γ-crystallin interacted with the membrane. However, at a Chol/POPC mixing ratio of 1, no association of γ-crystallin with the membrane was observed, which resulted in a defect-free membrane, and the severity of the membrane defect was decreased when βL-crystallin interacted with the membrane. The semi-transmembrane or transmembrane defects formed by the interaction of βL- and γ-crystallin on phospholipid membrane might be responsible for light scattering and cataract formation. However, Chol suppressed the formation of such defects in the membrane, likely maintaining lens membrane homeostasis and protecting against cataract formation

Understanding how children and young people with disabilities experience COVID-19 and humanitarian emergencies in low- and middle-income countries: a scoping review protocol

Introduction: People with disabilities have been disproportionately affected by the COVID-19 pandemic and there is a need to understand the diversity of experience globally to plan and implement responses. The experience of children and young people with disabilities is often overlooked as research focuses on younger children or adults, and they are not often engaged in the design of research methods to capture their experience. As part of a research project to develop participatory methods with children and young people with disabilities in rural Nepal to research their experience of COVID-19 (PARDAN), we sought to map the literature about experiences of children and young people with disabilities in emergency and COVID-19 contexts, and describe the methods used. This review will inform the development of our methods and will help identify gaps in the literature. / Methods: We will use Arksey and O’Malley’s (1) and Levac et al’s (2) framework to search for grey and published literature. We will search Scopus, Web of Science, ASSIA, Jstor, Source, Google Scholar, Academia.edu, Researchgate, and Google. We will also search websites of organisations working in emergencies such as United Nations High Commissioner for Refugees (UNHCR), World Health Organisation (WHO), United Nations Children's Fund (UNICEF), Humanity and Inclusion (HI), CBM International, International Disability Alliance (IDA), Women’s Refugee Commission (WRC), and National Federation of the Disabled Nepal (NFDN). We will approach our advisory committee and personal contacts in disabled persons organisations in the global south to help us locate additional relevant grey literature. Predetermined inclusion and exclusion criteria will be applied by five reviewers to determine study eligibility. Studies published in English and Nepali and between May 2011 and May 2021 will be reviewed. Methods and findings from literature will be extracted and a descriptive summary will be presented to an advisory committee of disabled persons organisations in Nepal. / Discussion: Through this review we will identify research gaps and develop implications of our findings which will be presented in a webinar and in publications. The review will be an important resource for those seeking to conduct research about children and young people with disabilities and will inform a global inclusive response to the pandemic which includes the perspectives of children and young people

Prevalence of intestinal parasitic infestations in school going children of Udaya Kharka Secondary school, Chapagaun, Lalitpur, Nepal

Intestinal Parasitic infestation is major cause of childhood morbidity in developing countries like Nepal with lack of hygiene and safe water supply. This study was done to find the actual scenario of parasitic infestation in a population of Nepal. This cross sectional, prospective study was undertaken in Udaya Kharkha Secoundary School, Chapagaun, Lalitpur. The study population included 246 students from school of age 6 years to 18 years. Routine and microscopic examination was done for stool collected from healthy students. Children among examined incidence of infestation was 52%. And different six types of parasites were found. Among which Giardiasis was the common with 42%. There was high incidence of parasitic infestations in school going children in the school. Incidence of parasitic infestations according to sex showed that girls are infested more than boys and according to age group 11 to 14years age group is highly infested

The Importance of Rating Scale Design in the Measurement of Patient-Reported Outcomes Using Questionnaires or Item Banks

This article is made available with the permission of the publisher, Association for Research in Vision and OphthalmologyPurpose.: To investigate the effect of rating scale designs (question formats and response categories) on item difficulty calibrations and assess the impact that rating scale differences have on overall vision-related activity limitation (VRAL) scores. Methods.: Sixteen existing patient-reported outcome instruments (PROs) suitable for cataract assessment, with different rating scales, were self-administered by patients on a cataract surgery waiting list. A total of 226 VRAL items from these PROs in their native rating scales were included in an item bank and calibrated using Rasch analysis. Fifteen item/content areas (e.g., reading newspapers) appearing in at least three different PROs were identified. Within each content area, item calibrations were compared and their range calculated. Similarly, five PROs having at least three items in common with the Visual Function (VF-14) were compared in terms of average item measures. Results.: A total of 614 patients (mean age ± SD, 74.1 ± 9.4 years) participated. Items with the same content varied in their calibration by as much as two logits; “reading the small print” had the largest range (1.99 logits) followed by “watching TV” (1.60). Compared with the VF-14 (0.00 logits), the rating scale of the Visual Disability Assessment (1.13 logits) produced the most difficult items and the Cataract Symptom Scale (0.24 logits) produced the least difficult items. The VRAL item bank was suboptimally targeted to the ability level of the participants (2.00 logits). Conclusions.: Rating scale designs have a significant effect on item calibrations. Therefore, constructing item banks from existing items in their native formats carries risks to face validity and transmission of problems inherent in existing instruments, such as poor targeting