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BackgroundThe 3xTg-AD transgenic mouse model of Alzheimer’s disease (AD) is an important tool to investigate the relationship between development of pathological amyloid-β (Aβ) and tau, neuroinflammation, and cognitive impairments. Traditional behavioral tasks assessing aspects of learning and memory, such as mazes requiring spatial navigation, unfortunately suffer from several shortcomings, including the stress of human handling and not probing species-typical behavior. The automated IntelliCage system was developed to circumvent such issues by testing mice in a social environment while measuring multiple aspects of cognition. Water consumption can serve as a primary motivator for task engagement. Once animals adapt to the cage and can access water, mice can be subjected to operant tasks. Each of the four corners of a cage contains doors to manipulate access to water, visual LED cues, and a valve allowing administration of an air puff. Previously, we detected significant impairments in 3xTg-AD mice in the IntelliCage, however a high failure rate and genotypical differences in water motivation were observed.MethodsHere, we implemented an IntelliCage paradigm where mice underwent progressively more difficult reaction time tasks to assess attention and impulsivity, behaviors mediated by the prefrontal cortex. Mice were placed in the IntelliCage at 11.5 months of age, which corresponds with the presence of widespread pathology.ResultsAs the difficulty of the reaction time tasks increased, 3xTg-AD mice exhibited lower percent Correct Responses than NonTg. When implementing varying pre-cue durations, where animals are required to wait between the initiation of the trial and the LED turning on (which then requires a nose-poke to access water), 3xTg-AD mice prematurely nose-poked on trials requiring a longer delay before a second nose poke would allow water access, demonstrating heightened impulsivity. The presence of soluble and insoluble fractions of cortical Aβ40 and 42, and phosphorylated tau epitopes threonine 181 and serine 396 confirmed the presence of neuropathological hallmarks in 3xTg-AD mice.ConclusionTogether, this study describes a novel protocol that overcomes motivational differences and detects attention and impulsivity deficits in 3xTg-AD mice utilizing the IntelliCage.

Background: During exercise, athletes experience a significant elevation in physiological strain, influenced by a multitude of internal and external factors, including physical and environmental conditions. These factors dictate how different organ systems respond to physical activity. This study aims to investigate the psychophysiological strain reactions to endurance training in military male runners. Methodology: A cohort of 75 male military runners, engaging in daily 90 minute exercise routines and aged between 20 and 30, was recruited for the study. The study assessed various physiological variables in military runners, encompassing lactate threshold (LT), maximum oxygen uptake (VO2 max), and percentage heart rate reserve (%HRR). Additionally, psychological assessments were conducted using the Brunel Mood Scale and Borg's Rate of Perceived Exertion. Results: The findings revealed a gradual increase in heart rate (HR) and VO2 max as running intensity escalated. Notably, the runners' perceived exertion rate rose following competitive running events, concomitant with a decrease in their positive mood scale ratings. This trend reflected their disappointment stemming from their performance outcomes. Conclusion: The study highlights that VO2 max and LT serve as robust indicators of male runners' potential. Furthermore, the evaluation of runners' mood and perception of exertion before and after the sprint demonstrated an augmentation in negative emotions post-exercise.

Abstract Background Skin, the most visible human entity, quickly displays emotional, physical, and psychological well-being. Research has shown a linear correlation between both types of skin aging within 30–69 years. Thus, the current research study aimed to highlight the use of SCINEXA, the novel skin aging score, in predicting accelerated biological aging. Methodology This cross-sectional study was conducted in Karachi, Pakistan, from 8th October 2021 to 4th December 2021. Both male and female subjects of age groups from 19 to 69 years were included. A novel skin aging score, 'SCINEXA' (Score of Intrinsic and Extrinsic Skin Aging), was used to assess skin aging. Both extrinsic and intrinsic signs were analyzed in terms of pigmented spots, coarse wrinkles, solar elastosis, telangiectasia, and laxity & seborrheic keratosis, respectively. Results The SCINEXA score indicated that 91.3% of participants had low aging signs and decreased pigmentation, while 2.9% had relatively high pigmentation on the forehead, cheek, forearm, and back of the hand. Coarse Wrinkles of grade 5 on the forehead were present among 19.7% of individuals, and 20.6% showed the same in the crow feet area. The skin aging symptoms are most significantly associated with age (p

Background:Obesity is an associated risk of devastating diseases and increased mortality. Thereby, the study objective was to evaluate the predisposing obese individuals towards accelerated ageing and determine the smell test as the possible indicator for early biological ageing.Methodology:This was a descriptive cross-sectional study carried out in Karachi, Pakistan, from 8thJune 2016-4thDecember 2019. A total of 412 individuals were included in the study. The data for age, gender, body mass index (BMI), sniffing score, and hemoglobin A1c(HbA1c)levels were collected from the target population. The screening 12 test was intended to be used for the examination of smell disorders in humans. The test results were used to differentiate between Normosmia, Hyposmia, and Anosmia. Results:Four hundred and twelve participants were recruited in the study with a mean age of 32.65±12.6 years and divided into three age groups; 20 to 36, 37 to 53, and54 to 70 years. There were 194 (47.1 %) females and 218 (52.9%) males. Out of the total, 19.9% of individuals were underweight, 58.5% were normal, 21.6% were overweight. It was also observed that of the total, 24% of participants were pre-diabetic. Cross analysis of the variables showed that the olfaction decreased as the BMI was increasing (p=0.000 for obese).Conclusion:As the sense of olfaction decreased with an increase in BMI and HbA1c levels, it was concluded that obese individuals have an increased predisposition towards accelerated ageing and the increased risk of predisposition to diabetes to obesity significantly accelerate the process of biological ageing.

Background: Chewing before performing a cognitive task increases oxygen levels in the Central Nervous System (CNS) areas important for processes of learning and memory. This study was done to evaluate and compare the effects of chewing gum on reaction time, visual short term memory, selective attention, verbal and non-verbal reasoning, and problem-solving ability in healthy male and female subjects. Methodology: The comparative, gender-based, interventional study was conducted involving 300 individuals placed in the control (n=150) and interventional group (n=150). Participants in the interventional group were required to chew gum till they completed the task. A questionnaire was designed to record the reaction time, memory, attention, executive and intellectual functioning and time took to solve each parameter. Each subject in both control and interventional group completed the questionnaire with and without chewing gum, respectively. Results for the two groups were compared using SPSS version 20.0. Results: It was found that the gum chewing group performs significantly better than the control group, thus chewing gum significantly improves cognitive performance. These cognitive effects of chewing were comparable among the two genders but relatively more pronounced among male participants as compared to females. Conclusion: Chewing gum is positively associated with higher level of cognitive performance than controls.

Background: Cortisol has a positive correlation with Obesity, which indicates a relation between obesity and cortisol. Furthermore, there are increased chances of demolished muscle strength up to two times with elevated levels of cortisol. Thus, the aim of the study was to assess the impact of increased cortisol levels on the grip strength and to determine that the grip strength is a possible indicator for accelerated biological ageing. Methodology: This cross-sectional, purposive sampling study was conducted in Karachi, Pakistan. Overall 412 were recruited to participate in the study, the data for age, gender, BMI, grip strength, and cortisol was collected from the participants. The age-wise correlates of hand grip strength were assessed using Multiple linear regression analysis and the mean decline in the grip strength with advancing age is displayed. Results: The mean Grip strength of the overall population was 30.75±14.35. It was observed that there was a negative significant correlation between the age and the grip strength (F = 3.57; r= -0.27; p=0.007) i.e. with increasing age a gradual decrease in the grip strength was observed. BMI (Obesity) was also significantly associated with grip strength (p=0.000). Moreover, females were more likely to have weak muscle strength as compared to males (p=0.000). However, no significant association was observed among the grip strength, socioeconomic status & longevity in any of the studied age groups (p>0.05). While a significant correlation was found between Grip strength and Obesity as well as between Obesity and Cortisol. Conclusion: Thus it can be concluded that obese individuals had increased chances of accelerated biological ageing due to an increase in cortisol, and a weakened grip strength is an indicator of it.

Background: The foremost factor involved in Neurodegeneration is the impact of epigenetic modifications; through its nature to epigenetically mark the neuron-associated genes, also, by affecting cognitive functions and damaging neurons that promote mutations. Due to these changes in the genes; neurodegenerative diseases are developed. This review will assess epigenetic modifications that switch “on” & “off” the genes associated with neurons that lead towards neurodegeneration in humans. Methodology: This systematic review is based on Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) guidelines to conduct a search strategy and for the preparation of the manuscript. A search engine (PubMed) was used and the article reference list was searched for relevant primary research articles. 100 out of 22278 studies dated from January/2000 to February/2019 met the inclusion criteria. Two quality assessments were piloted and included: (1) Authors evaluation and (2) Risk of bias. Results: Quality of interventions provided was rated “good”, Risk of bias in studies was rated “fair” and the team of authors approved included papers. Furthermore, 13 out of 100 studies critical appraisal analysis demonstrated the relationship between epigenetic alterations and neurodegeneration and the rest of the studies described neuro-epigenetics, epigenetic remodeling and epigenetic mechanisms. Conclusion: Exogenous influence like aviation stress or co-factors, such as nutrition and physical stress plays a major role in silencing the “gene switching” proteins of epigenetic marks and influences the onset and progression of neurodegeneration. Furthermore, intervention in epigenetics might help promote brain health.

Background: The energy drink (ED) consumption rapidly increased by youngsters in recent years. It has a high concentration of sugar, caffeine, taurine and other stimulants that enhance the mental and physical activity. The aim of the current study was to evaluate the comparative effects of taurine-based energy drinks on blood glucose (BG) level and Prothrombin time (PT) among healthy male and female subjects. Methodology: A cross-sectional single center observational study was conducted from over a sample of 50 subjects between 18 to 25 years of age. The subjects were kept in two distinct groups as males and females and all assessments were made individually for both genders. Written informed consent was taken from each subject prior to enrolment in the study. As per the study protocol, the BG level (mg/dl) and PT (sec) were taken twice. Each subject was asked to drink 250 ml taurine based carbonated energy drink after taking the first recording of both (BG level and PT). And secondary records are taken 1 hour after the consumption of ED. The collected data was statistically analyzed using SPSS version 16. Results: The mean age of the study subjects was 22.34±2.3 years. It was seen that the post-test mean BG level significantly increased to 129.2±18.3 mg/dl (males) and 147.92±24.4 mg/dl (females). Moreover, PT was decreased both in males (96±45.8 sec) and females (78.6±20.2 sec) after ED consumption. Conclusion: ED consumption contributed to the increased BG level and increased coagulation (decreased PT) and hence indicating an increased risk of thrombosis and type 2 diabetes among persistent consumers. Further large-scale studies are required locally in order to provide sufficient evidence.

Background: Music is beneficial for adults, it enhances psychological comfort, self-confidence, self-esteem, and lowers the symptoms of anxiety, pain and depression, relieving stress by improving life quality and thus providing satisfaction. It holds the capacity to initiate a multitude of cognitive processes in the brain. We aim to evaluate and compare the effects of music on reaction time, attention, working capacity of short-term memory and verbal fluency of male and female subjects. Methodology: An observational study was conducted on a sample of 300 subjects either males or females between the age groups of 19 to 30 years, affiliated with different universities and institutes. After inclusion, these subjects were divided into two groups, standard and experimental group with 150 subjects in each group. Ruler & Drop method test and Stroop test with or without music were used to test the reaction time (RT) and attention, respectively. Whereas, working capacity of short-term memory was tested using the George A. Miller rule of memory. And verbal fluency was evaluated using semantic verbal fluency (SVF) and phonological verbal fluency (PVF) tests. The data was analyzed using SPSS Version 22. Results: The comparative mean values between the groups for RT, attention, short-term memory and verbal fluency scores were greater among the subjects in the experimental group as the tests were performed in association with musical interference. Mean reaction time for both visual and tactile cues were significantly increased in the experimental group i.e. 0.151±0.034 (males) and 0.124 ±0.050 (females) for tactile cues and 0.150 ±0.042 (males) and 0.152 ±0.033 (females). Moreover, Stroop interference also increased in both genders while short-term memory score declined from high to average and verbal fluency was also compromised due to musical interference. Conclusion: It is concluded from the study results that music holds both positive and negative effects on brain activity. It imparts positive effect on both RT and attention but in case of the working capacity of short-term memory and verbal fluency, the effects are negative.

The Arabic word for health صحي is synonymous in English language to “weight” وزن (Khoury, 2001), and a cultural link for perceiving weight as health can very well be understood, the impact of Arabic culture and values on a random Pakistani cannot be denied under any circumstances. Obesity is a major concern, in adults and more so in children. Pakistan is going through a transitional phase for some decades whereby changes are coming across in lifestyles, family structures, incomes, a new set of stresses, rural to urban shift, migrants’ influx making the transition all the more complex when compared to other countries (Nanan, 2002).

Background: Aroma or scent therapy is one of the traditional treatments, which is presumed to be helpful in supporting and balancing the mind, body and soul. It is a complementary therapy that has been clinically proven to be beneficial in improving various attributes of life. The aim of the study was to evaluate the effectiveness of aromatherapy on the physiological vitals of the healthy females. Methodology: An observational study has been conducted to determine the effectiveness of Aromatherapy on Vitals and Physical Factors. This study targeted to recruit a small set of healthy females for the study. All the readings were collected at room temperature, between the months of November 2015 to January 2016. The recordings were compared by the two-tailed test, with probability taken as 1, to detect the influence of aromatherapy on physiological workings of the healthy human body that has been exposed to the physical strain due to the moderate level of exercise. The test involved the comparative analysis of the physiological vitals of those females who exercised regularly; their vitals were evaluated after a course of physical activity in two sets i.e. with and without aromatherapy. Results: The result showed a significant difference in the pre and post status mainly in the readings for the cardiac output, pulse rate and body temperature, increases after physical activity of average intensity and is further supported by the aromatherapy. Conclusion: The study came to the conclusion that aromatherapy has beneficial influences on the rate of body metabolism.

Cognition is an intellective action of understanding and gaining knowledge through speculation, acquaintance and practices that result in awareness, impression and perception. Cognition, Proprioception and balance all are aspects of ability of the brain to perform at high level at expense of endurance, speed and power. However, increasing age is linked with decrements in periodic memory. Thus, exercise positively impacts brain utility during middle age, particularly frontal lobe-mediated intellectual processes, like organization, programing, reticence and operational memory. The aim of the present study was to observe whether exercise improve cognitive skills or not. The study was conducted on 200 healthy male subjects equally divided in two groups: exercising males and non-exercising males. The subjects were asked to fill in a proforma consisting of visual test and a reading test, which aimed at comparative evaluation of the duration of short term memory between exercisers and non-exercisers. It was found that exercise impacts on frontal lobe to enhance intellectual processes and hippocampus memory processing.

Tea is the most widely consumed beverage after water around the globe with consumer-base of both young and old, while energy drinks are popular primarily among young adults. Among the different varieties available black tea is the most commonly used and contains L-Theanine, theobromine, theophylline and the highest amount of caffeine. Energy drinks on the other hand contain taurine, herbs, B vitamins and stimulant drugs chiefly caffeine. The purpose of the study conducted was to evaluate the effect of regular tea and energy drinks on attention. A survey was conducted and subjects were given a task that was designed to test the above mentioned parameter and time taken to solve each part of the task was accurately noted. Results of the energy drink group and the tea group were compared to the control group. Gender based differences were also analyzed. Our results lead us to conclude that the most widely taken beverages enhance attention in both genders, with women outperforming men.

Regular exercise is largely done to have maintained body shape and fitness. It exerts influence on almost all physiological and psychological parameters. One of these is dilutional anemia or sports anemia that is the initial adaptation of body to exercise. The aim of present study was to evaluate the extent of dilutional anemia in healthy males who have been exercising for past three years at least. Randomly selected 40 male subjects from gymnasiums of Karachi were recruited in the study. Peripheral blood was collected pre and post exercise, and tested for hematocrit, hemoglobin concentration and red blood cell count. Increased hemoglobin concentration was observed which is directly related to increased red blood cell count which is inversely proportional to blood plasma level. So, decreased plasma level is the possible cause that pseudo dilution anemia was not observed in males who have been regularly exercising for past three years.

Exercising in gym is largely aimed at getting a desirably muscled physique that is subject to peer approval as well as associated with a higher self-esteem. Many people opt for workouts that define the lean body mass, whereas others aim at getting a beefed up body. The latter being a long and tiring process, pushes many people to opt for power enhancing drugs, for boosting their athletic endurance. Among others; steroids, erythropoietin, creatine, protein supplements are available over the counter all over Pakistani Pharmacies. Together with increasing the muscle bulk, they exert highly undesirable effects on hematological as well as lipid parameters. The present study evaluated and compared the side effects reported by the users of such PED on their mind, personality and body. A proforma was filled by each subject. The results showed that depending on the type of PED used, it has altered the personality of the individual significantly. Attributes that their personality and body did not have initially were observed by them upon chronic usage of these ergogenic aids. This lead us to conclude that “amount in PRs” is not the only price these PED users pay for attaining their desired beefed up physique, with which emerged another question, is that cost actually worth it?

The use of genetically diverse mouse models offers a more accurate reflection of human genetic variability, improving the translatability of findings to heterogeneous human populations. This approach is particularly valuable in understanding diverse immune responses to disease by environmental exposures. This study investigates the inflammatory responses to acute exposures to mainstream cigarette smoke (CS) and environmental tobacco smoke (ETS) in two genetically diverse mouse strains, CC002/UncJ (UNC) & Diversity Outbred (J:DO). The UM-HET3 (HET3) mouse strain, typically used in aging intervention studies, has also been used to evaluate the translatability of this model for age-associated pathologies. The study involves a comprehensive approach, including BALF cytokine analysis, evaluation of lung tissue architecture, assessment of macrophages and its associated proteins (MMP9 & MMP12) abundance. Several cytokines/chemokines were found to be upregulated across three strains. Notably, the UNC strain exclusively showed upregulation of TNF-α, IL-17A, and IL-13, whereas the J:DO showed an upregulation in KC. The number of alveolar macrophages in the lungs of UNC mice was very low at baseline compared to other strains studied in this study, which is indicative of some inherent shift in the pulmonary immune profiles of these inbred mice. In contrast, the J:DO strain, characterized by genetic outbreeding, showed a much more robust lung macrophage response comparable to C57BL/6J. The findings provide valuable insight into how genetic diversity affects immune responses in response to acute CS/ETS exposure, with implications for understanding diverse human responses to environmental stressors in studying lung pathophysiology., (© 2025 The Author(s). Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

Background: Transient receptor potential vanilloid type 4 (TRPV4) is a versatile ion channel with diverse roles in immune cells, including macrophages. While its function in inflammation remains debated, we investigated its role in regulating IL-10 production and its impact on macrophage reprogramming during inflammation., Methods: We investigated the connection between TRPV4 activation and CREB-mediated IL-10 production during inflammation. Notably, this signaling pathway was found to reprogram macrophages and enhance their ability to resist inflammatory damage. The experiments were conducted on primary macrophages and were further corroborated by animal studies., Results: In response to TRPV4 activation during inflammation, we observed a significant increase in intracellular Ca 2+ levels, which triggered the activation of the transcription factor CREB, subsequently upregulating IL-10 production. This IL-10 played a pivotal role in reprogramming macrophages to withstand inflammatory damage. Using a mouse model of acute lung injury (ALI), we confirmed that TRPV4 activation during ALI led to IL-10 secretion, but this increase did not significantly contribute to inflammation resolution. Moreover, we found that TRPV4 prevented the accumulation of dysfunctional mitochondria in macrophages through the CREB-IL-10 axis during inflammation. Suppression of CREB or TRPV4 inhibition exacerbated mitochondrial dysfunction, while treatment with recombinant IL-10 mitigated these effects. Additionally, IL-10 induced mitophagy and cleared dysfunctional mitochondria in LPS-exposed cells., Conclusion: Our study highlights the essential role of TRPV4 in regulating IL-10 production and mitochondrial health in macrophages during inflammation. These findings contribute to understand the role of TRPV4 in immune responses and suggest potential therapeutic targets for modulating inflammation-induced cellular dysfunction., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Oral cancer is a global health challenge with a difficult histopathological diagnosis. The accurate histopathological interpretation of oral cancer tissue samples remains difficult. However, early diagnosis is very challenging due to a lack of experienced pathologists and inter- observer variability in diagnosis. The application of artificial intelligence (deep learning algorithms) for oral cancer histology images is very promising for rapid diagnosis. However, it requires a quality annotated dataset to build AI models. We present ORCHID (ORal Cancer Histology Image Database), a specialized database generated to advance research in AI-based histology image analytics of oral cancer and precancer. The ORCHID database is an extensive multicenter collection of high-resolution images captured at 1000X effective magnification (100X objective lens), encapsulating various oral cancer and precancer categories, such as oral submucous fibrosis (OSMF) and oral squamous cell carcinoma (OSCC). Additionally, it also contains grade-level sub-classifications for OSCC, such as well- differentiated (WD), moderately-differentiated (MD), and poorly-differentiated (PD). The database seeks to aid in developing innovative artificial intelligence-based rapid diagnostics for OSMF and OSCC, along with subtypes., (© 2024. The Author(s).)

Idiopathic Pulmonary Fibrosis (IPF) is a progressively fatal and incurable disease characterized by the loss of alveolar structures, increased epithelial-mesenchymal transition (EMT), and aberrant tissue repair. In this study, we investigated the role of Nuclear Factor I-B (NFIB), a transcription factor critical for lung development and maturation, in IPF. Using both human lung tissue samples from patients with IPF, and a mouse model of lung fibrosis induced by bleomycin, we showed that there was a significant reduction of NFIB both in the lungs of patients and mice with IPF. Furthermore, our in vitro experiments using cultured human lung cells demonstrated that the loss of NFIB was associated with the induction of EMT by transforming growth factor beta (TGF-β). Knockdown of NFIB promoted EMT, while overexpression of NFIB suppressed EMT and attenuated the severity of bleomycin-induced lung fibrosis in mice. Mechanistically, we identified post-translational regulation of NFIB by miR-326, a miRNA with anti-fibrotic effects that is diminished in IPF. Specifically, we showed that miR-326 stabilized and increased the expression of NFIB through its 3'UTR target sites for Human antigen R (HuR). Moreover, treatment of mice with either NFIB plasmid or miR-326 reversed airway collagen deposition and fibrosis. In conclusion, our study emphasizes the critical role of NFIB in lung development and maturation, and its reduction in IPF leading to EMT and loss of alveolar structures. Our study highlights the potential of miR-326 as a therapeutic intervention for IPF. The schema shows the role of NFIB in maintaining the normal epithelial cell characteristics in the lungs and how its reduction leads to a shift towards mesenchymal cell-like features and pulmonary fibrosis. A In normal lungs, NFIB is expressed abundantly in the epithelial cells, which helps in maintaining their shape, cell polarity and adhesion molecules. However, when the lungs are exposed to factors that induce pulmonary fibrosis, such as bleomycin, or TGF-β, the epithelial cells undergo epithelial to mesenchymal transition (EMT), which leads to a decrease in NFIB. B The mesenchymal cells that arise from EMT appear as spindle-shaped with loss of cell junctions, increased cell migration, loss of polarity and expression of markers associated with mesenchymal cells/fibroblasts. C We designed a therapeutic approach that involves exogenous administration of NFIB in the form of overexpression plasmid or microRNA-326. This therapeutic approach decreases the mesenchymal cell phenotype and restores the epithelial cell phenotype, thus preventing the development or progression of pulmonary fibrosis., (© 2023. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Mesenchymal stem cell (MSC) transplantation alleviates metabolic defects in diseased recipient cells by intercellular mitochondrial transport (IMT). However, the effect of host metabolic conditions on IMT and thereby on the therapeutic efficacy of MSCs has largely remained unexplored. Here we found impaired mitophagy, and reduced IMT in MSCs derived from high-fat diet (HFD)-induced obese mouse (MSC-Ob). MSC-Ob failed to sequester their damaged mitochondria into LC3-dependent autophagosomes due to decrease in mitochondrial cardiolipin content, which we propose as a putative mitophagy receptor for LC3 in MSCs. Functionally, MSC-Ob exhibited diminished potential to rescue mitochondrial dysfunction and cell death in stress-induced airway epithelial cells. Pharmacological modulation of MSCs enhanced cardiolipin-dependent mitophagy and restored their IMT ability to airway epithelial cells. Therapeutically, these modulated MSCs attenuated features of allergic airway inflammation (AAI) in two independent mouse models by restoring healthy IMT. However, unmodulated MSC-Ob failed to do so. Notably, in human (h)MSCs, induced metabolic stress associated impaired cardiolipin-dependent mitophagy was restored upon pharmacological modulation. In summary, we have provided the first comprehensive molecular understanding of impaired mitophagy in obese-derived MSCs and highlight the importance of pharmacological modulation of these cells for therapeutic intervention. A MSCs obtained from (HFD)-induced obese mice (MSC-Ob) show underlying mitochondrial dysfunction with a concomitant decrease in cardiolipin content. These changes prevent LC3-cardiolipin interaction, thereby reducing dysfunctional mitochondria sequestration into LC3-autophagosomes and thus impaired mitophagy. The impaired mitophagy is associated with reduced intercellular mitochondrial transport (IMT) via tunneling nanotubes (TNTs) between MSC-Ob and epithelial cells in co-culture or in vivo. B Pyrroloquinoline quinone (PQQ) modulation in MSC-Ob restores mitochondrial health, cardiolipin content, and thereby sequestration of depolarized mitochondria into the autophagosomes to alleviate impaired mitophagy. Concomitantly, MSC-Ob shows restoration of mitochondrial health upon PQQ treatment (MSC-ObPQQ). During co-culture with epithelial cells or transplantation in vivo into the mice lungs, MSC-ObPQQ restores IMT and prevents epithelial cell death. C Upon transplantation in two independent allergic airway inflammatory mouse models, MSC-Ob failed to rescue the airway inflammation, hyperactivity, metabolic changes in epithelial cells. D PQQ modulated MSCs restored these metabolic defects and restored lung physiology and airway remodeling parameters., (© 2023. The Author(s).)

Circulating cell-free mitochondrial DNA (cf-mtDNA) has been found in the plasma of severely ill COVID-19 patients and is now known as a strong predictor of mortality. However, the underlying mechanism of mtDNA release is unexplored. Here, we show a novel mechanism of SARS-CoV-2-mediated pro-inflammatory/pro-apoptotic mtDNA release and a rational therapeutic stem cell-based approach to mitigate these effects. We systematically screened the effects of 29 SARS-CoV-2 proteins on mitochondrial damage and cell death and found that NSP4 and ORF9b caused extensive mitochondrial structural changes, outer membrane macropore formation, and the release of inner membrane vesicles loaded with mtDNA. The macropore-forming ability of NSP4 was mediated through its interaction with BCL2 antagonist/killer (BAK), whereas ORF9b was found to inhibit the anti-apoptotic member of the BCL2 family protein myeloid cell leukemia-1 (MCL1) and induce inner membrane vesicle formation containing mtDNA. Knockdown of BAK and/or overexpression of MCL1 significantly reversed SARS-CoV-2-mediated mitochondrial damage. Therapeutically, we engineered human mesenchymal stem cells (MSCs) with a simultaneous knockdown of BAK and overexpression of MCL1 (MSCshBAK+MCL1) and named these cells IMAT-MSCs (intercellular mitochondrial transfer-assisted therapeutic MSCs). Upon co-culture with SARS-CoV-2-infected or NSP4/ORF9b-transduced airway epithelial cells, IMAT-MSCs displayed functional intercellular mitochondrial transfer (IMT) via tunneling nanotubes (TNTs). The mitochondrial donation by IMAT-MSCs attenuated the pro-inflammatory and pro-apoptotic mtDNA release from co-cultured epithelial cells. Our findings thus provide a new mechanistic basis for SARS-CoV-2-induced cell death and a novel therapeutic approach to engineering MSCs for the treatment of COVID-19.

To discover anticancer drugs with novel structures and expand our research scope, pyrazoline derivatives ( 3a - 3l ) were designed and synthesized through cyclization of chalcones with thiosemicarbazide/semicarbazide in CH 3 COOH as a solvent. All newly synthesized pyrazoline derivatives were fully characterized using several spectroscopic experiments such as 1 H, 13 C NMR, FT-IR spectroscopy, and mass analysis. By HPLC , the purity of all analogs was found above 95% and both lead compounds ( 3a and 3h ) were also validated by HRMS . Anticancer activity of synthesized pyrazoline derivatives ( 3a - 3l ) was investigated by the MTT assay against the human lung cancer cell (A549), human cervical cancer cell (HeLa), and human primary normal lung cells (HFL-1). Staurosporine (STS) was used as a standard drug. The anticancer results showed that two potent analogs 3a and 3h exhibit excellent activity against A549 (IC 50 = 13.49 ± 0.17 and 22.54 ± 0.25 μM) and HeLa cells (IC 50 = 17.52 ± 0.09 and 24.14 ± 0.86 μM) and low toxicity against the HFL-1 (IC 50 = 114.50 ± 0.01 and 173.20 ± 10 μM). The flow cytometry was further used to confirm the anticancer activity of potent derivatives against the A549 cancer cell line. DNA binding interaction of anticancer agents 3a and 3h with Ct-DNA has been carried out by absorption, fluorescence, EtBr (dye displacement assay), circular dichroism, cyclic voltammetry and time-resolved fluorescence, which showed noncovalent binding mode of interaction. Anticancer activity of both lead compounds ( 3a and 3h ) may be attributed to DNA binding. The evaluation of the antioxidant potential of pyrazoline analogs 3a and 3h by 2,2-diphenyl-1-picrylhydrazyl free radical showed promising antioxidant activity with IC 50 values of 0.132 ± 0.012 and 0.215 ± 0.025 μg/mL, respectively. In silico molecular docking of pyrazoline derivatives was also performed using autodock vina software against the DNA hexamer with PDB ID: 1Z3F and ADMET properties to explore their best hits., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

The global burden of disease caused by a respiratory syncytial virus (RSV) is becoming more widely recognized in young children and adults. Heparan sulfate helps in attaching the virion through G protein with the host cell membrane. In this study, we examined the structural changes of ectodomain G protein (edG) in a wide pH range. The absorbance results revealed that protein maintains its tertiary structure at physiological and highly acidic and alkaline pH. However, visible aggregation of protein was observed in mild acidic pH. The intrinsic fluorescence study shows no significant change in the λ max except at pH 12.0. The ANS fluorescence of edG at pH 2.0 and 3.0 forms an acid-induced molten globule-like state. The denaturation transition curve monitored by fluorescence spectroscopy revealed that urea and GdmCl induced denaturation native (N) ↔ denatured (D) state follows a two-state process. The fluorescence quenching, molecular docking, and 50 ns simulation measurements suggested that heparan sulfate showed excellent binding affinity to edG. Our binding study provides a preliminary insight into the interaction of edG to the host cell membrane via heparan sulfate. This binding can be inhibited using experimental approaches at the molecular level leading to the prevention of effective host-pathogen interaction.

A major bottleneck in scaling-up COVID-19 testing is the need for sophisticated instruments and well-trained healthcare professionals, which are already overwhelmed due to the pandemic. Moreover, the high-sensitive SARS-CoV-2 diagnostics are contingent on an RNA extraction step, which, in turn, is restricted by constraints in the supply chain. Here, we present CASSPIT ( C as13 A ssisted S aliva-based & S martphone I ntegrated T esting), which will allow direct use of saliva samples without the need for an extra RNA extraction step for SARS-CoV-2 detection. CASSPIT utilizes CRISPR-Cas13a based SARS-CoV-2 RNA detection, and lateral-flow assay (LFA) readout of the test results. The sample preparation workflow includes an optimized chemical treatment and heat inactivation method, which, when applied to COVID-19 clinical samples, showed a 97% positive agreement with the RNA extraction method. With CASSPIT, LFA based visual limit of detection (LoD) for a given SARS-CoV-2 RNA spiked into the saliva samples was ~200 copies; image analysis-based quantification further improved the analytical sensitivity to ~100 copies. Upon validation of clinical sensitivity on RNA extraction-free saliva samples (n = 76), a 98% agreement between the lateral-flow readout and RT-qPCR data was found (Ct<35). To enable user-friendly test results with provision for data storage and online consultation, we subsequently integrated lateral-flow strips with a smartphone application. We believe CASSPIT will eliminate our reliance on RT-qPCR by providing comparable sensitivity and will be a step toward establishing nucleic acid-based point-of-care (POC) testing for COVID-19., Competing Interests: Author RB is employed by 360 Diagnostic and Health Services, author DJG is employed by Noodle Analytics Pvt Ltd. and author GDJ is employed by Valerian Chem Pvt. Ltd. A patent for the saliva-based and RNA extraction free detection of SARS-CoV2 using CRISPR Diagnostics has been filed. TA, MJ, JI, GJ, and RK are the inventors and applicants in the patent. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Azmi, Faizan, Kumar, Raj Yadav, Chaudhary, Kumar Singh, Butola, Ganotra, Datt Joshi, Deep Jhingan, Iqbal, Joshi and Ahmad.)

The classical necroptosis signaling is mediated by death receptors (DRs) that work in synergy with traditional caspase inhibitory signals. Currently, potential therapeutic molecules are in various phases of clinical trials for a spectrum of pathological conditions associated with necroptosis. However, a non-classical model of necroptosis has also emerged over the last decade with a relatively unexplored molecular mechanism. Although in vitro studies and preclinical models have shown its close association with mitochondrial dysfunction (mito-dysfunction), contradictory reports have emerged which complicate its definitiveness. Though impaired mitochondrial calcium ([Ca 2+ ] m ) handling is established in necrotic cell death, how this interplay regulates necroptosis is yet to be elucidated. Taking these questions into consideration, we have discussed various molecular aspects of necroptosis with the emerging role of mito-dysfunction. Based on the central role of altered [Ca 2+ ] m handling in mito-dysfunction mediated necroptosis, we have provided a comprehensive molecular insight into this emerging paradigm. Potential reasons for the contradictory findings regarding the role of mito-dysfunction in necroptosis in general and mitochondrial-dependent necroptosis in specific are discussed. We also provide insights into the current understanding of how [Ca 2+ ] m can be a critical determinant in deciding the cell fate under certain pathological conditions, while under others it may be dispensable. Lastly, we have highlighted the key molecular targets which have a direct implication for therapeutic intervention in conditions that are associated with impaired [Ca 2+ ] m handling and cell death by necroptosis., (Copyright © 2020 Elsevier B.V. and Mitochondria Research Society. All rights reserved.)

N-formyl pyrazoline derivatives (3a-3l) were designed and synthesized via Michael addition reaction through cyclization of chalcones with hydrazine hydrate in presence of formic acid. The structural elucidation of N-formyl pyrazoline derivatives was carried out by various spectroscopic techniques such as 1 H, 13 C NMR, FT-IR, UV-visible spectroscopy, mass spectrometry and elemental analysis. Anticancer activity of the pyrazoline derivatives (3a-3l) was evaluated against human lung cancer (A549), fibrosarcoma cell lines (HT1080) and human primary normal lung cells (HFL-1) by MTT assay. The results of anticancer activity showed that potent analogs 3b and 3d exhibited promising activity against A549 (IC 50  = 12.47 ± 1.08 and 14.46 ± 2.76 µM) and HT1080 (IC 50  = 11.40 ± 0.66 and 23.74 ± 13.30 µM) but low toxic against the HFL-1 (IC 50  = 116.47 ± 43.38 and 152.36 ± 22.18 µM). The anticancer activity of potent derivatives (3b and 3d) against A549 cancer cell line was further confirmed by flow cytometry based approach. DNA binding interactions of the pyrazoline derivatives 3b and 3d have been carried out with calf thymus DNA (Ct-DNA) using absorption, fluorescence and viscosity measurements, circular dichroism and cyclic voltammetry. Antioxidant potential of N-formyl pyrazoline derivatives (3a-3l) has been also estimated through DPPH (2,2-diphenyl-1-picrylhydrazyl) free radical and H 2 O 2. Results revealed that all the compounds exhibited significant antioxidant activity. In silico molecular modelling and ADMET properties of pyrazoline derivatives were also studied using PyRx software against topoisomerase II receptor with PDB ID: 1ZXM to explore their best hits. MD simulation of 3b and 3d was also carried out with topoisomerase II for structure-function correlation in a protein. HuTopoII inhibitory activity of the analogs (3a-3l) was examined by relaxation assay at varying concentrations 100-1000 µM., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: Oral submucous fibrosis (OSMF) is a premalignant condition mainly caused by areca nut chewing and is characterized by progressive fibrosis of submucosal tissues and epithelial atrophy. Activation of transforming growth factor beta (TGF-β) signaling is considered main causative event for increased collagen production and fibrosis. In this study, molecular pathogenesis of OSMF was investigated based on the expression of the TGF-β genes in OSMF tissues compared to normal controls., Methods: A total of 33 OSMF and 10 normal tissues were collected from patients and their clinic-epidemiological data was recorded. The expression of TGF-β isoform genes- TGF β1, TGF β2, TGF β3 and its receptor TGF βR1, TGF βR2 was studied by real time polymerase chain reaction (PCR). Comparison of the expression of these genes among normal controls and OSMF patients was done. The PCR results were confirmed by histopathological and immunohistochemical staining., Results: The histological changes included atrophic epithelium, loss of rete ridges, presence of inflammatory cells and dense collagen bundles in connective tissue. PCR showed statistically significant upregulation of TGF-β isoforms in OSMF as compared to normal tissues. Of the three isoforms, maximum fold change was observed in TGF-β1. Similarly, both TGF-βR1 and TGF-βR2 were found to be elevated in OSMF tissues compared to normal. The semi-quantitative analysis by immunohistochemical staining revealed statistically significant difference between normal and OSMF tissues., Conclusion: TGF-β signaling plays a major role in the molecular pathogenesis of OSMF as shown by increased mRNA expression of all the three TGF-β isotypes and their receptors., Competing Interests: None declared., (© 2020 Craniofacial Research Foundation. Published by Elsevier B.V. All rights reserved.)

Chikungunya virus (CHIKV) is a re-emerging pathogen of global importance. We attempted to gain an insight into the organisation, distribution and mutational load of the virus strains reported from different parts of the world. We describe transmission dynamics and genetic characterisation of CHIKV across the globe during the last 65 years from 1952 to 2017. The evolutionary pattern of CHIKV was analysed using the E1 protein gene through phylogenetic, Bayesian and Network methods with a dataset of 265 sequences from various countries. The time to most recent common ancestor of the virus was estimated to be 491 years ago with an evolutionary rate of 2.78 × 10-4 substitutions/site/year. Genetic characterisation of CHIKV strains was carried out in terms of variable sites, selection pressure and epitope mapping. The neutral selection pressure on the E1 gene of the virus suggested a stochastic process of evolution. We identified six potential epitope peptides in the E1 protein showing substantial interaction with human MHC-I and MHC-II alleles. The present study augments global epidemiological and population dynamics of CHIKV warranting undertaking of appropriate control measures. The identification of epitopic peptides can be useful in the development of epitope-based vaccine strategies against this re-emerging viral pathogen.

Background: Dengue is a rapidly emerging arthropod borne viral infection affecting tropical and sub-tropical regions of the world. Dengue is an acute febrile illness but sometimes causes more fatal complications like dengue haemorrhagic fever (DHF) and dengue shock syndrome (DSS). Delhi, the capital of India has become hyper endemic for dengue virus because all the four serotypes are circulating here., Methods: The present study describes the identification of dengue virus from clinical samples collected from the suspected dengue patients from New Delhi, India during 2016. The CprM region of Dengue virus genome was analyzed for phylogenetic, selection pressure and Shannon entropy analyses., Results: The present study reports circulation of a single serotype (DENV-3) in New Delhi, during 2016. The phylogenetic analysis revealed that Indian subcontinent (genotype III) of DENV-3 was circulating in Delhi during this period. Neutral selection pressure in the analyzed region revealed relatively conserved nature of this part of the Dengue virus genome. Amino acid at 31 was positively selected and had high entropy value suggesting probability of variation at this position., Conclusions: The changing trend in circulation of dengue virus serotypes necessitates the continuous epidemiological surveillance for the dengue outbreaks in this region., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Co-circulation of Chikungunya and Dengue viral infections (CHIKV and DENV) have been reported mainly due to transmission by common Aedes vector. The purpose of the study was to identify and characterise the circulating strains of CHIKV and DENV in DENV endemic region of New Delhi during 2016. CHIKV and DENV were identified in the blood samples (n = 130) collected from suspected patients by RT-PCR. CHIKV was identified in 26 of 65 samples (40%). Similarly, DENV was detected in 48 of 120 samples (40%). Co-infection with both the viruses was identified in five (9%) of the samples. Interestingly, concurrent infection with DENV, CHIKV and Plasmodium vivax was detected in two samples. CHIKV strains (n = 11) belonged to the ECSA genotype whereas DENV-3 sequences (n = eight) clustered in Genotype III by phylogenetic analysis. Selection pressure of E1 protein of CHIKV and CprM protein of DENV-3 revealed purifying selection with four and two positive sites, respectively. Four amino acids of the CHIKV were positively selected and had high entropy suggesting probable variations. Co-circulation of both viruses in DENV endemic regions warrants effective monitoring of these emerging pathogens via comprehensive surveillance for implementation of effective control measures.

Zika virus is an arthropod-borne re-emerging pathogen associated with the global pandemic of 2015-2016. The devastating effect of Zika viral infection is reflected by its neurological manifestations such as microcephaly in newborns. This scenario evoked our interest to uncover the neurotropic localization, multiplication of the virus, and the mechanism of microcephaly. The present report provides an overview of a possible molecular mechanism of Zika virus-induced microcephaly based on recent publications. Transplacental transmission of Zika viral infection from mother to foetus during the first trimester of pregnancy results in propagation of the virus in human neural progenitor cells (hNPCs), where entry is facilitated by the receptor (AXL protein) leading to the alteration of signalling and immune pathways in host cells. Further modification of the viral-induced TLR3-mediated immune network in the infected hNPCs affects viral replication. Downregulation of neurogenesis and upregulation of apoptosis in hNPCs leads to cell cycle arrest and death of the developing neurons. In addition, it is likely that the environmental, physiological, immunological, and genetic factors that determine in utero transmission of Zika virus are also involved in neurotropism. Despite the global concern regarding the Zika-mediated epidemic, the precise molecular mechanism of neuropathogenesis remains elusive., (© 2017 S. Karger AG, Basel.)