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Extracellular Vesicles (EVs) have gained high repute in drug delivery systems owing to their relatively higher efficacy as natural drug delivery vehicles. The current literature has advanced the use of EVs in drug delivery through exploring various aspects including their biogenesis, characterization, and nanoengineering techniques thereby leading them from laboratory to clinical use with optimized good laboratory practices. In this timely review, we summarize the current status of EVs characterizations and recent updates on nanoengineering of EVs regarding Cargo loading and surface fabrication. Further, we have also reviewed current progress in clinical translation and implications of EVs in clinical trials together with future recommendations.

Chimeric antigen receptor (CAR) T cell treatment is an emerging subject following its curative response in hematological metastasis. However, solid tumors present a number of obstructions which have been a bull's eye to steer the CARs toward another victory in solid tumor microenvironment (TME). To combat against solid tumors, the construction, transfection and delivery of CARs is obliged to nano-engineering for better results and success in clinical trials. Herein, in this minireview, we discuss some of the potential and novel applications of nanotechnology to engineer better performing CARs to target solid TME. Moreover, we highlight potential gaps and strategies to overcome for future advancements in nano immunoengineering.

Cell-free DNA (cfDNA) are un-encapsulated DNA fragments present in biological fluids ranging in an average size of up to 200 base pairs. The novel use of cfDNA is a prime candidate in the diagnostic and prognostic approach to unveiling many inflammatory diseases, especially cancer. Moreover, their potential as biomarkers is due to their ubiquitous presence in the body, non-invasive nature, and aiding in a different autopsy method. This review will focus on the diagnostic and prognostic potential of cfDNA as non-invasive biomarkers in oncology.

Epilepsy affects millions of people worldwide. Although antiepileptic drugs work for the majority of epileptic patients, these drugs do not work for some of the patients, subjecting them to drug-resistant epilepsy (DRE). Voltage-gated sodium channels act as targets for a number of antiepileptic drugs, and the genes encoding these channels can play a crucial role in developing drug-resistant epilepsy. This case-control (100 control: 101patients) study evaluated the association of sodium channel genes SCN1A and SCN2A with drug-resistant epilepsy. The cases were further accounted in two categories, drug-resistant and drug-responsive epileptic patients. The polymorphic sites rs794726754, rs1057518252, rs121918809, rs12191792, rs121917932, c.730 G > T, c.735 G > T, c.736 A > T, rs10167228, and rs2298771 of the SCN1A gene and rs17183814 of SCN2A gene were selected for mutational analysis. The DNA was isolated, amplified by PCR, and then, was run through 1% agarose gel. The sequencing was performed, and the sequences were observed through BioEdit software for any change in DNA sequence. In our study, no polymorphism was observed in the studied SNPs except for rs2298771. For rs2298771, a significant difference existed in the distribution of genotypic and allelic frequencies (p 0.01). Our study indicated that the rs2298771 polymorphism of SCN1A may not be associated with chance of developing DRE in the Pakistani population.

Epilepsy is a major global issue. Epilepsy patients are treated with AED (antiepileptic drugs). Interindividual variability in drug response has been documented in several studies. The resistance to drug response may be attributed to genetic polymorphism. The current study was undertaken to investigate the CYP2C9 gene polymorphism associated with antiepileptic drug (AED) resistance in the Pakistani population. The current study included 337 individuals including 100 control subjects, 110 drug-resistant subjects, and 127 drug responders. Genomic DNA was isolated from blood, and amplification of rs1799853 (430C > T) and rs1057910 was carried out by polymerase chain reaction. Genotypes of CYP2C9 SNPs were determined by Sanger’s sequencing. Astounding results were observed in the current study that none of the well-known reported SNPs of CYP2C9 was found in our Pakistani cohorts. However, a novel missense variant (c.374G > A) was found only in drug-resistant patients of the current study. According to the in silico analysis performed by PolyPhen-2, it was observed that this nonsynonymous substitution is likely to be pathogenic. The results of our study demonstrated that rs1799853 and rs1057910 may be involved in drug resistance in the Pakistani population. However, some other variants on CYP2C9 may play a critical role in AED resistance that needs to be explored.

Arthritis is a genetic disorder characterized by bones and joint degradation assisted by severe pain and inflammation. It is evident by the studies that 0 candidate genes variations play vital role in its development and progression. Therefore, we investigated the genetic variation of TLR-8, TNF, and ESR-1α genes in the Pakistani population. A case-control study comprising 300 RA, 316 OA, and 412 control subjects was conducted. PCR-RFLP and direct sequencing methods were used for determining genetic variations. Analysis was performed by using PLINK and MEGA 6.0 software. Allelic and genetic frequencies of polymorphisms identified on rs3764879 (TLR-8), rs3764880 (TLR-8), rs5744080 (TLR-8), rs1800629 (TNF), rs2228480 (ESR-1α), and rs1451501590 (ESR-1α) were significantly varied among RA, OA, and controls. Novel functional mutations SCV000844945 and SCV000844946 on TLR-8 as well as a non-functional SCV000804801 and functional variation SCV000804802 on ESR-1α were also identified and reported for the first time in the studied population. Multiple site analyses indicated that polymorphisms on TLR-8 and ESR-1α genes were significant risk factors in disease onset to the next generation. In conclusion, TLR-08 and ESR-1α were significant in the onset of arthritis whereas the TNF was not found as a significant risk factor in the onset of RA and OA.

The present study investigated the (i) socio-demographic predictors of psychological distress, (ii) socio-demographic predictors of satisfaction from online classes, and (iii) the relationship between psychological distress and satisfaction from online classes among university students of Pakistan during the COVID-19 pandemic. An online questionnaire-based survey was conducted. A total of 2220 respondents that was enrolled at the University of the Punjab (PU), University of Management and Technology (UMT), and the University of Central Punjab (UCP) were involved in the current study. Data were collected at a 64% response rate and analyzed with SPSS IBM Version 21.0. Results revealed that approximately 41% of the students were facing severe psychological distress while about 65% were found unsatisfied with online classes. Besides, a linear negative relationship between the independent variable, i.e. psychological distress and the dependent variable, i.e. satisfaction from online classes was found. Therefore, to minimize the level of psychological distress and increase students’ satisfaction with online classes it is highly recommended to take precautionary measures by the relevant stakeholders.

BPA, a ubiquitously used plasticizer, has become one of the contaminants of emerging concern and causes many serious health implications in humans due to multiple exposure pathways. The current study was aimed at investigating the deformities of structures that arise by exposure of the small intestine to BPA through trace elements estimation of tissues as well as the study of serum profile. Two major groups of Wistar rats were established: one control group and the other experimental group, which was further divided into four groups based on dose (10 mg/kg/bodyweight and 25 mg/kg/bodyweight, respectively) and duration of exposure (6 and 12 weeks, respectively). Histological study of the small intestine showed the distorted structures in the experimental groups. The special staining performed illustrated the accumulation of calcium deposits in the small intestinal tissue in treated groups. Trace metals estimation showed a significant increase in the metallic content of sodium and iron and a decrease in the calcium content in the experimental groups (p=0.05). Serum profiling illustrated an increase in total iron-binding capacity and glucose levels and a decrease in the serum total iron level (p=0.05). An increased expression of a proinflammatory cytokine (IFN-α) was observed in the liver. From all these findings, it was inferred that BPA caused many structural alterations in the small intestinal tissue, which further affected its functioning. The calcium deposits seen through special staining affected the motility of the small intestine and caused its dysfunction. It was also induced from serum profiling that BPA affected the homeostasis of iron and glucose and caused its imbalance. Also, as BPA got absorbed from the small intestine and reached the liver via the blood stream, it caused hepatoxicity in the liver and led to increased inflammatory response by IFN-α against the toxicant.

The process of paper production requires a huge quantity of water and energy and in turn contributes a number of effluents in the form of phenolics, toxic organic compounds and heavy metals in wastewater (leachate). The present investigation was aimed to assess the toxic effects of leachate on liver micro-architecture and heavy metal elements of the liver. Eighteen (18) healthy male Wistar rats (240 ± 10g) were selected and acclimatized prior to experimental treatment. These rats were randomly divided into three groups viz, Control group (received 4ml/ kg normal saline), Group 1 (4ml/ kg leachate) and Group 2 (4ml/ kg 1:10 diluted leachate). All the animals were dissected and liver tissues were collected and processed accordingly after 24 h of leachate treatment. High level of cadmium and chromium were found in Group 1 as compared to the control group upon liver metal contents analysis found out by flame atomic absorption spectrometer. A clear disruption of micro-architecture of the liver, congested sinusoids, damaged central vein, and perturbed morphology was observed in Group-1 as revealed by H & E staining. Moreover, loss of polarity, congestion, and disruption of hepatocytes and pronounced vacuolization in the cytoplasm was observed in Group 2 compared to control sections. On the basis of above findings, it can be concluded that paper industry leachate is highly toxic and its intraperitoneal injection results in hepatotoxicity that not only affects the hepatic micro-architecture but also results in perturbed liver metal contents. Therefore, proper treatment of such wastewater is required before its disposal.

Genetic polymorphismsof MYOCalter the myocilin protein,which leads to disruption of thenormal regulation of intraocular pressure (IOP) that ultimately causes glaucoma.Theaim of the present study was to identify the polymorphism in exon 3 of the MYOC gene of theglaucoma patients in Lahore, Pakistan. We conducted a case-control study with 100 patients and 100 controls subjects. We extracted DNA from blood samples,amplified the target DNA fragmentby PCR, and identifiedpolymorphisms through sequencing. We observed that the allelic and genotypic frequencies of rs74315341 and rs879255525 were associated with glaucoma in our patient population. The polymorphism atrs74315341 led to the substitutionof serine for arginine,whereas the polymorphism at rs879255525 led to the substitution ofasparagine for lysine. The haplotype TGAAGCCATTTC was associated with disease onset, whereas the haplotype GGAAGCCATTTC was protective against disease development. In conclusion, weidentified MYOC gene polymorphisms in susceptible regions that were associated withglaucoma onset among the Lahore patient population.This is the first report to identify a novel mutation in rs879255525 in exon 3 of the MYOC genethat is associated withglaucoma.

This study was aimed to find histological changes in the extrahepatic organs, hepatic iron deposition, and gene expression of some iron regulatory proteins in rats after sterile muscle abscess during the acute intoxication of Nerium oleander leaves decoction. 10 ml/kg of the leaves extract was injected intramuscularly in Wistar rats (200–225 g, n=4). Control animals received saline injection of matched volume. Animals were anesthetized and sacrificed after 3, 6, 12, and 24 h after administration of decoction. Lungs, kidney, spleen, and liver were extracted and processed for histopathological examination while portion of liver tissue was proceeded for iron regulatory gene expression quantification. Sections of all studied organs were found with signs of cellular dysfunction with infiltration of variety of leucocytes. In the lungs section at 3 h time point mononuclear cell infiltrates were observed while in alveolar tissue at 24 h time point dilation and even collapse in some of the alveoli were evident. In kidney sections distortion of renal tubules and epithelial cells with shrinkage of glomeruli was noted at all studied time points. In the splenic section of 12 h time point, degeneration, depopulation, and shrinkage of white pulp have been noted. Distension of the red pulp along with activation of splenic follicles was evident after 24 h onset of APR. Significant changes in the expression of acute phase cytokine and iron regulatory genes were noted. IL-6 and Hepc gene expression were strongly upregulated up to 12 h whereby Tf gene expression showed an early upregulation at 3 h time point followed by downregulation on later points while Hjv gene expression showed an overall downregulation at all study time points compared to control. It is concluded that inherent toxins present in the N. oleander can induce acute phase response and cause severe histological changes in the organs and marked changes in the regulation of iron regulatory proteins thus cannot be practiced routinely.

Fagonia indica is a traditionally used phytomedicine to cure hepatic ailments. However, efficient validation of its hepatoprotective effect and molecular mechanisms involved are not yet well established. Therefore, the present study was designed to evaluate the hepatoprotective activity of Fagonia indica and to understand the molecular mechanisms involved in the reversal of hepatic injury. The liver injury mouse model was established by thioacetamide followed by oral administration of plant extract. Serum biochemical and histological analyses were performed to assess the level of hepatic injury. Expression analysis of proinflammatory, hepatic, and immune regulatory genes was performed with RT-PCR. Results of serological and histological analyses described the restoration of normal liver function and architecture in mice treated with plant extract. In addition, altered expression of proinflammatory (IL-1β, IL-6, TNF-α, and TGF-β) and hepatic (krt-18 and albumin) markers further strengthens the liver injury reversal effects of Fagonia indica. Furthermore, a significant expression regulation of innate immunity components such as toll-like receptors 4 and 9 and MyD-88 was observed suggesting an immune regulatory role of the plant in curing liver injury. In conclusion, the current study not only proposes Fagonia indica, a strong hepatoprotective candidate, but also recommends an immune regulatory toll-like receptor pathway as an important therapeutic target in liver diseases.

Hepatocellular carcinoma is one of the fatal malignancies and is considered as the third leading cause of death. Mutations, genetic modifications, dietary aflatoxins, or impairments in the regulation of oncogenic pathways may bring about liver cancer. An effective barrier against hepatotoxins is offered by gut-liver axis as a change in gut permeability and expanded translocation of lipopolysaccharides triggers the activation of Toll-like receptors which stimulate the process of hepatocarcinogenesis. Prebiotics, nondigestible oligosaccharides, have a pivotal role to play when it comes to inducing an antitumor effect. A healthy gut flora balance is imperative to downregulation of inflammatory cytokines and reducing lipopolysaccharides induced endotoxemia, thus inducing the antitumor effect.

Processing of fossil fuels is the major environmental issue today. Biomass utilization for the production of chemicals presents an alternative to simple energy generation by burning. Lignocellulosic biomass (cellulose, hemicellulose and lignin) is abundant and has been used for variety of purposes. Among them, lignin polymer having phenyl-propanoid subunits linked together either through C-C bonds or ether linkages can produce chemicals. It can be depolymerized by fungi using their enzyme machinery (laccases and peroxidases). Both acetic acid and formic acid production by certain fungi contribute significantly to lignin depolymerization. Fungal natural organic acids production is thought to have many key roles in nature depending upon the type of fungi producing them. Biological conversion of lignocellulosic biomass is beneficial over physiochemical processes. Laccases, copper containing proteins oxidize a broad spectrum of inorganic as well as organic compounds but most specifically phenolic compounds by radical catalyzed mechanism. Similarly, lignin peroxidases (LiP), heme containing proteins perform a vital part in oxidizing a wide variety of aromatic compounds with H2O2. Lignin depolymerization yields value-added compounds, the important ones are aromatics and phenols as well as certain polymers like polyurethane and carbon fibers. Thus, this review will provide a concept that biological modifications of lignin using acidophilic fungi can generate certain value added and environmentally friendly chemicals.

Pakistan is among the last three countries along with Afghanistan and Nigeria, where polio virus is still endemic. More or less, with some fluctuations, numbers of reported cases in the past few years have shown a rising trend. Year 2014 pushed the country into the deep sea of difficulties, as number of cases rose to red alert level of 328. Security situation has adversely affected the whole immunization coverage campaign. In a country where 40 polio vaccinators have been killed since 2012, such a big number of cases is not a surprising outcome. Worse perception of parents about polio vaccine as in Karachi and FATA, the high risk zones, makes 100% coverage a dream. Minor and perhaps delayed payments to polio workers make them frustrated, resulting in decline of trained manpower for vaccination. Strong implementation of policies is required and those found guilty of attack on polio workers need to be punished. Targeted community awareness programme, strong surveillance network, and involvement of influential religious entities can help to root out polio disease from country. Present review is aimed at analyzing all barriers on the road to success in eradication of polio from Pakistan.

Interleukin-22 (IL-22) is a pluripotent T cell-derived cytokine which is a member of IL-10 cytokine family. It is the only interleukin produced by immune cells but does not target immune system components. IL-22 is mainly produced by dendritic cells (DCs) and TH17, TH22, NK, and NKT cells and targets a number of body tissues including liver, pancreas, and other epithelial tissues. It provokes a series of downstream signaling pathways upon binding with IL-22R complex which protects liver damage through STAT3 activation. IL-22BP is an inhibitor of IL-22 which has 20–1000x more affinity to bind with IL-22 compared to IL-22R1 that inhibits IL-22 activity. Its level was found to be positively correlated with the severity of liver damage and fibrosis. So, the present review is an effort to reveal the exact mechanism lying in the hepatoprotective activity of IL-22 and some of its future therapeutic implications.

A vast variety of nonstructural proteins have been studied for their key roles and involvement in a number of biological phenomenona. Interleukin-32 is a novel cytokine whose presence has been confirmed in most of the mammals except rodents. The IL-32 gene was identified on human chromosome 16 p13.3. The gene has eight exons and nine splice variants, namely, IL-32α, IL-32β, IL-32γ, IL-32δ, IL-32ε, IL-32ζ, IL-32η, IL-32θ, and IL-32s. It was found to induce the expression of various inflammatory cytokines including TNF-α, IL-6, and IL-1β as well as macrophage inflammatory protein-2 (MIP-2) and has been reported previously to be involved in the pathogenesis and progression of a number of inflammatory disorders, namely, inflammatory bowel disease (IBD), gastric inflammation and cancer, rheumatoid arthritis, and chronic obstructive pulmonary disease (COPD). In the current review, we have highlighted the involvement of IL-32 in gastric cancer, gastric inflammation, and chronic rhinosinusitis. We have also tried to explore various mechanisms suspected to induce the expression of this extraordinary cytokine as well as various mechanisms of action employed by IL-32 during the mediation and progression of the above said problems.

Cypermethrin, a type II pyrethroid, is one of the most widely used insecticides in Pakistan. It is considered to be a safe pesticide; however, the possible health hazards of this pyrethroid have been ignored. Cypermethrin may become an air pollutant and adversely affect the health of non-target organisms, leading to acute or chronic disorders. The present work aims to investigate the effects of cypermethrin on lung and liver tissues due to inhalation exposure. The study is performed on 16 mature Swiss albino mice, including controls. The animals are divided into 4 groups (4 mice each) and exposed to 0.5 % dilution of cypermethrin in an inhalation chamber (40×35×25 cm3) for different time periods, whereas control animals are not exposed to any insecticide. The histopathological changes in lungs and liver tissues reveal that cypermethrin exposure induces time dependent changes in the liver and in the lungs. It damages the normal organization of liver tissues, causing liver injury due to necrosis, significant decrease in number of cells, and widening of sinusoids and fibrosis. Inhalation exposure of cypermethrin results in significant hyperplasia, clumping of cells and necrosis in the lungs. It also induces pulmonary edema, alveolitis, and pulmonary fibrosis by the deposition of collagen. Taking these findings together, it may be concluded that cypermethrin and other pyrethroids cause hazardous effects in non-target organisms through inhalation exposure. Serious efforts and awareness are required to monitor and reduce the insecticide induced health hazards in third world countries. doi:10.14456/WJST.2014.67