The Secret Life of lncRNAs: Conserved, yet Not Conserved.

Guo and colleagues discover a new layer of complexity to the lncRNA evolution where positionally conserved lncRNAs in human ESCs are broadly spliced and exported to the cytoplasm contrary to their mouse counterpart that are predominantly unspliced and nuclear retained. Distinct processing leads to species-specific lncRNA function in pluripotency maintenance.

Graphene Hybrid Tough Hydrogels with Nanostructures for Tissue Regeneration.

Over the past few decades, hydrogels have attracted considerable attention as promising biomedical materials. However, conventional hydrogels require improved mechanical properties, such as brittleness, which significantly limits their widespread use. Recently, hydrogels with remarkably improved toughness have been developed; however, their low biocompatibility must be addressed. In this study, we developed a tough graphene hybrid hydrogel with nanostructures. The resultant hydrogel exhibited remarkable mechanical properties while representing an aligned nanostructure that resembled the extracellular matrix of soft tissue. Owing to the synergistic effect of the topographical properties, and the enhanced biochemical properties, the graphene hybrid hydrogel had excellent stretchability, resilience, toughness, and biocompatibility. Furthermore, the hydrogel displayed outstanding tissue regeneration capabilities (e.g., skin and tendons). Overall, the proposed graphene hybrid tough hydrogel may provide significant insights into the application of tough hydrogels in tissue regeneration.

Micro/nanoengineered agricultural by-products for biomedical and environmental applications.

Agriculturally derived by-products generated during the growth cycles of living organisms as secondary products have attracted increasing interest due to their wide range of biomedical and environmental applications. These by-products are considered promising candidates because of their unique characteristics including chemical stability, profound biocompatibility and offering a green approach by producing the least impact on the environment. Recently, micro/nanoengineering based techniques play a significant role in upgrading their utility, by controlling their structural integrity and promoting their functions at a micro and nano scale. Specifically, they can be used for biomedical applications such as tissue regeneration, drug delivery, disease diagnosis, as well as environmental applications such as filtration, bioenergy production, and the detection of environmental pollutants. This review highlights the diverse role of micro/nano-engineering techniques when applied on agricultural by-products with intriguing properties and upscaling their wide range of applications across the biomedical and environmental fields. Finally, we outline the future prospects and remarkable potential that these agricultural by-products hold in establishing a new era in the realms of biomedical science and environmental research.

Graphene Hybrid Inner Ear Organoid with Enhanced Maturity.

Inner ear organoids (IEOs) are 3D structures grown in vitro, which can mimic the complex cellular structure and function of the inner ear. IEOs are potential solutions to problems related to inner ear development, disease modeling, and drug delivery. However, current approaches in generating IEOs using chemical factors have a few limitations, resulting in unpredictable outcomes. In this study, we propose the use of nanomaterial-based approaches, specifically by using graphene oxide (GO). GO's unique properties promote cell-extracellular matrix interactions and cell-cell gap junctions, thereby enhancing hair cell formation, which is an essential part of IEO development. We also investigated the potential applications for drug testing. Our findings suggest that GO is a promising candidate for enhancing the functionality of IEOs and advancing our understanding of the problems underlying inner ear development. The use of nanomaterial-based approaches may provide a more reliable and effective method for building better IEOs in the future.

Chloroquine induces transitory attenuation of proliferation of human lung cancer cells through regulation of mutant P53 and YAP.

BACKGROUND: Non-small cell lung carcinoma (NSCLC) is the most common cause of cancer-associated deaths worldwide. Though recent development in targeted therapy has improved NSCLC prognosis, yet there is an unmet need to identify novel causative factors and appropriate therapeutic regimen against NSCLCs. METHODS AND RESULTS: In this study, we identify key molecular factors de-regulated in NSCLCs. Analyze their expression by real-time PCR and immunoblot; map their localization by immuno-fluorescence microscopy. We further propose an FDA approved drug, chloroquine (CQ) that affects the function of the molecular factors and hence can be repurposed as a therapeutic strategy against NSCLCs. Available NSCLC mutation data reflects a high probabilistic chance of patients harboring a p53 mutation, especially a gain of function (GOF)-R273H mutation. The GOF-P53 mutation enables the P53 protein to potentially interact with non-canonical protein partners facilitating oncogenesis. In this context, analysis of existing transcriptomic data from R273H-P53 expressing cells shows a concomitant up-regulation of Yes-associated protein (YAP) transcriptional targets and its protein partner TEAD1 in NSCLCs, suggesting a possible link between R273H-P53 and YAP. We therefore explored the inter-dependence of R273H-P53 and YAP in NSCLC cells. They were found to co-operatively regulate NSCLC proliferation. Genetic or pharmacological inhibition of YAP and GOF-P53 resulted in sensitization of NSCLC cells. Further analysis of pathways controlled by GOF-P53 and YAP showed that they positively regulate the cellular homeostatic process- autophagy to mediate survival. We hence postulated that a modulation of autophagy might be a potent strategy to curb proliferation. In accordance to above, autophagy inhibition, especially with the FDA-approved drug- chloroquine (CQ) resulted in cytoplasmic accumulation and reduced transcriptional activity of GOF-P53 and YAP, leading to growth arrest of NSCLC cells. CONCLUSION: Our study highlights the importance of GOF-P53 and YAP in NSCLC proliferation and proposes autophagy inhibition as an efficient strategy to attenuate NSCLC tumorigenesis.

Clinicopathologic characteristics and outcomes of late onset lupus nephritis: a single centre experience.

Systemic Lupus Erythematosus (SLE) occurs in the reproductive age group. Renal involvement occurs less frequently in late-onset SLE than in reproductive-age SLE patients. Here, we aimed to study the clinical, serological and histopathological characteristics of late-onset lupus nephritis (LN). Late-onset LN was defined as disease onset after 47 years of age, corresponding to the average menopausal age. Records of biopsy proven late-onset lupus nephritis patients diagnosed between June 2000 and June 2020 were reviewed. Late-onset LN constituted 53 of 4420 patients (1.2%) biopsied during the study period. Females represented 90.65% of the cohort. Mean age of the cohort was 49.5 ± 7.05 years at the time of SLE diagnosis while its renal presentation was delayed by median duration of 10 months (IQR 3-48 months). Renal failure was present in 28 patients (52.8%) with acute kidney injury (AKI) (28.3%, n = 15) as the most common presentation. On histopathological analysis, class IV was observed in 23 patients (43.5%), crescents were observed in one-third cases and lupus vasculopathy in 4 patients (7.5%). All patients received steroids. Majority of patients (43.3%; n = 23) received Euro lupus protocol for induction. On median follow up duration of 82 months, renal flares were noted in 9 patients (17%) and 8 patients (15.1%) became dialysis dependent. Among 11 patients (21%) with infectious complications, 7 patients (13.2%) suffered from tuberculosis. Infections caused three-fourth of the deaths. Late-onset lupus nephritis is rare and presents as renal failure in majority. Renal biopsy affects the clinical decision of judicious use of immunosuppression which is imperative due to high rate of infections in this cohort.

Dynamic alterations of H3K4me3 and H3K27me3 at ADAM17 and Jagged-1 gene promoters cause an inflammatory switch of endothelial cells.

Histone protein modifications control the inflammatory state of many immune cells. However, how dynamic alteration in histone methylation causes endothelial inflammation and apoptosis is not clearly understood. To examine this, we explored two contrasting histone methylations; an activating histone H3 lysine 4 trimethylation (H3K4me3) and a repressive histone H3 lysine 27 trimethylation (H3K27me3) in endothelial cells (EC) undergoing inflammation. Through computer-aided reconstruction and 3D printing of the human coronary artery, we developed a unique model where EC were exposed to a pattern of oscillatory/disturbed flow as similar to in vivo conditions. Upon induction of endothelial inflammation, we detected a significant rise in H3K4me3 caused by an increase in the expression of SET1/COMPASS family of H3K4 methyltransferases, including MLL1, MLL2, and SET1B. In contrast, EC undergoing inflammation exhibited truncated H3K27me3 level engendered by EZH2 cytosolic translocation through threonine 367 phosphorylation and an increase in the expression of histone demethylating enzyme JMJD3 and UTX. Additionally, many SET1/COMPASS family of proteins, including MLL1 (C), MLL2, and WDR5, were associated with either UTX or JMJD3 or both and such association was elevated in EC upon exposure to inflammatory stimuli. Dynamic enrichment of H3K4me3 and loss of H3K27me3 at Notch-associated gene promoters caused ADAM17 and Jagged-1 derepression and abrupt Notch activation. Conversely, either reducing H3K4me3 or increasing H3K27me3 in EC undergoing inflammation attenuated Notch activation, endothelial inflammation, and apoptosis. Together, these findings indicate that dynamic chromatin modifications may cause an inflammatory and apoptotic switch of EC and that epigenetic reprogramming can potentially improve outcomes in endothelial inflammation-associated cardiovascular diseases.

High mortality and residual kidney damage with Coronavirus disease-19-associated acute kidney injury in northern India.

BACKGROUND: Acute kidney injury (AKI) is associated with morbidity and mortality in COVID-19 patients. The incidence of AKI and its outcomes vary in different parts of the world. We aimed to analyze the AKI incidence, predictors of AKI, mortality, and renal function outcomes on follow-up in hospitalized patients with COVID-19. MATERIALS AND METHODS: The study was designed as a retrospective, observational study of electronically captured data on the hospital information system of laboratory-confirmed COVID-19 patients, with and without AKI, between March 2020 to June 2021. The predictor of AKI and mortality and residual damage in recovered AKI patients were analyzed. RESULTS: Of the 3395 patients, 3010 COVID-19 patients were eligible. AKI occurred in 951 (31.5%); with stages 1, 2, and 3 in 605 (63.7%), 138 (14.5%), and 208 (21.8%) patients, respectively. AKI severity increased with COVID-19 severity. Of 951 AKI patients, 403 died, and 548 were discharged. AKI group had higher mortality (42.3%) than the non-AKI (6.6%). At discharge, complete recovery was noticed in 370(67.5%), while 178 (32.5%) had residual damage. At three months of follow-up, 108 (69.6%) of 155 patients showed complete recovery. Residual damage was observed in 47 (30.3%). In 14 (9%) patients, serum creatinine remained elevated above the baseline. Thirty-three (21.2%) patients showed proteinuria (n = 24) and microscopic hematuria (n = 9). CONCLUSIONS: AKI is common among patients hospitalized with COVID-19 and is associated with high mortality. Residual kidney damage post-COVID-19 in recovered AKI patients may increase the CKD burden.

An NMR-based approach reveals the core structure of the functional domain of SINEUP lncRNAs.

Long non-coding RNAs (lncRNAs) are attracting widespread attention for their emerging regulatory, transcriptional, epigenetic, structural and various other functions. Comprehensive transcriptome analysis has revealed that retrotransposon elements (REs) are transcribed and enriched in lncRNA sequences. However, the functions of lncRNAs and the molecular roles of the embedded REs are largely unknown. The secondary and tertiary structures of lncRNAs and their embedded REs are likely to have essential functional roles, but experimental determination and reliable computational prediction of large RNA structures have been extremely challenging. We report here the nuclear magnetic resonance (NMR)-based secondary structure determination of the 167-nt inverted short interspersed nuclear element (SINE) B2, which is embedded in antisense Uchl1 lncRNA and upregulates the translation of sense Uchl1 mRNAs. By using NMR 'fingerprints' as a sensitive probe in the domain survey, we successfully divided the full-length inverted SINE B2 into minimal units made of two discrete structured domains and one dynamic domain without altering their original structures after careful boundary adjustments. This approach allowed us to identify a structured domain in nucleotides 31-119 of the inverted SINE B2. This approach will be applicable to determining the structures of other regulatory lncRNAs.

SINEUP long non-coding RNA acts via PTBP1 and HNRNPK to promote translational initiation assemblies.

SINEUPs are long non-coding RNAs (lncRNAs) that contain a SINE element, and which up-regulate the translation of target mRNA. They have been studied in a wide range of applications, as both biological and therapeutic tools, although the underpinning molecular mechanism is unclear. Here, we focused on the sub-cellular distribution of target mRNAs and SINEUP RNAs, performing co-transfection of expression vectors for these transcripts into human embryonic kidney cells (HEK293T/17), to investigate the network of translational regulation. The results showed that co-localization of target mRNAs and SINEUP RNAs in the cytoplasm was a key phenomenon. We identified PTBP1 and HNRNPK as essential RNA binding proteins. These proteins contributed to SINEUP RNA sub-cellular distribution and to assembly of translational initiation complexes, leading to enhanced target mRNA translation. These findings will promote a better understanding of the mechanisms employed by regulatory RNAs implicated in efficient protein translation.

Verteporfin disrupts multiple steps of autophagy and regulates p53 to sensitize osteosarcoma cells.

BACKGROUND: Osteosarcoma (OS) is a malignant tumor of the bone mostly observed in children and adolescents. The current treatment approach includes neoadjuvant and adjuvant chemotherapy; however, drug resistance often hinders therapy in OS patients. Also, the post-relapse survival of OS patients is as low as 20%. We therefore planned to understand the molecular cause for its poor prognosis and design an appropriate therapeutic strategy to combat the disease. METHODS: We analyzed OS patient dataset from Gene Expression Omnibus (GEO) and identified the differentially expressed genes and the top deregulated pathways in OS. Subsequently, drugs targeting the major de-regulated pathways were selected and the following assays were conducted- MTT assay to assess cytotoxicity of drugs in OS cells; immunoblotting and immunostaining to analyze key protein expression and localization after drug treatment; LysoTracker staining to monitor lysosomes; Acridine Orange to label acidic vesicles; and DCFDA to measure Reactive Oxygen Species (ROS). RESULTS: The differential gene expression analysis from OS patient dataset implicated the striking involvement of cellular processes linked to autophagy and protein processing in the development of OS. We therefore selected the FDA approved drugs, chloroquine (CQ) and verteporfin (VP) known for autophagy inhibitory and proteotoxic functions to explore against OS. Importantly, VP, but not CQ, showed an extensive dose-dependent cytotoxicity. It resulted in autophagy disruption at multiple steps extending from perturbation of early autophagic processes, inhibition of autophagic flux to induction of lysosomal instability. Interestingly, VP treated protein lysates showed a ROS-dependent high molecular weight (HMW) band when probed for P62 and P53 protein. Further, VP triggered accumulation of ubiquitinated proteins as well. Since VP had a pronounced disruptive effect on cellular protein homeostasis, we explored the possibility of simultaneous inhibition of the ubiquitin-proteasomal system (UPS) by MG-132 (MG). Addition of a proteasomal inhibitor significantly aggravated VP induced cytotoxicity. MG co-treatment also led to selective targeting of P53 to the lysosomes. CONCLUSION: Herein, we propose VP and MG induce regulation of autophagy and protein homeostasis which can be exploited as an effective therapeutic strategy against osteosarcoma.

Phenolic Extract of Seagrass, Halophila ovalis Activates Intrinsic Pathway of Apoptosis in Human Breast Cancer (MCF-7) Cells.

The marine ecosystem is considered as a treasure of numerous novel biologically active molecules. We investigated the anticancer potential of the phenolic extract of Halophila ovalis in breast cancer (MCF-7) cells and characterized the possible underlying molecular mechanism. The phenolic extract (5 µl) of H. ovalis effectively inhibited the growth of MCF-7 cells. The results of DAPI staining indicated that this phenolic extract potently induces apoptosis in MCF-7 cells which was observed by increased chromatin condensation in the treated cells. An increased expression of the active fragments of an executioner caspase, caspase 3 in phenolic extract-treated MCF-7 cells further confirms this apoptosis induction. In consequence, the loss of mitochondrial membrane potential was noticed in treated cells. The protein expression analyzes show decreased expression of the anti-apoptotic protein, Bcl-2, and DNA repair enzyme, PARP in treated cells indicating the probable molecular targets of apoptosis. Further, the phenolic extract of H. ovalis blocked the antioxidant defense system in MCF-7 cells by down-regulating the protein expression of a major transcription factor, Nrf-2 and regulatory antioxidant enzymes, SOD-2 and HO-1. These results show the presence of chemopreventive compound(s) in the phenolic extract, which offers a platform for future studies to identify the active principles.

Transcriptomic analysis associated with reversal of cisplatin sensitivity in drug resistant osteosarcoma cells after a drug holiday.

BACKGROUND: Resistance to chemotherapy is one of the major hurdles in current cancer therapy. With the increasing occurrence of drug resistance, a paradigm shift in treatment strategy is required. Recently "medication vacation" has emerged as a unique, yet uncomplicated strategy in which withdrawal of drug pressure for certain duration allowed tumor cells to regain sensitivity to the drug. However, little is known about the molecular alterations associated with such an outcome. METHODS: In this study, human osteosarcoma (OS) cells resistant to the extensively used drug cisplatin, were withdrawn from drug pressure, and thereafter cytotoxic response of the cells to the drug was evaluated. We further performed next-generation RNA sequencing and compared transcriptome between parental (OS), resistant (OS-R) and the drug withdrawn (OS-DW) cells. Differentially expressed transcripts were identified, and biological association network (BAN), gene ontology (GO) and pathway enrichment analysis of the differentially regulated transcripts were performed to identify key events associated with withdrawal of drug pressure. RESULTS: Following drug withdrawal, the sensitivity of the cells to the drug was found to be regained. Analysis of the expression profile showed that key genes like, IRAK3, IL6ST, RELA, AKT1, FKBP1A and ADIPOQ went significantly down in OS-DW cells when compared to OS-R. Also, genes involved in Wnt signaling, PI3K-Akt, Notch signaling, and ABC transporters were drastically down-regulated in OS-DW cells compared to OS-R. Although, a very small subset of genes maintained similar expression pattern between OS, OS-R and OS-DW, nonetheless majority of the transcriptomic pattern of OS-DW was distinctively different and unique in comparison to either the drug sensitive OS or drug resistant OS-R cells. CONCLUSION: Our data suggests that though drug withdrawal causes reversal of sensitivity, the transcriptomic pattern does not necessarily show significant match with resistant or parental control cells. We strongly believe that exploration of the molecular basis of drug holiday might facilitate additional potential alternative treatment options for aggressive and resistant cancers.

Botryomycosis: A surprising revelation in the lacrimal sac.

A middle aged woman presented to us with a localised well defined swelling of 3 months duration. It was located just below the lower eyelid punctum and was constantly discharging whitish granules. We suspected it to be arising from the lacrimal apparatus and posted the patient for Dacryocystectomy. On the operating table we found a swelling in the region of the lacrimal sac which was later excised. Histopathology revealed Botryomycosis and Chronic Dacryocystitis. Botryomycosis is a rare condition and requires a high index of suspicion to diagnose it. It is confirmed by histopathology and culture. Surgical debridement is the treatment of choice in such cases with an assessment of the immune status. Long term antibiotic treatment is required in all conditions as recurrence is common.

Establishment of a reborn MMV-microarray technology: realization of microbiome analysis and other hitherto inaccessible technologies.

BACKGROUND: With the accelerating development of bioscience, the problem of research cost has become important. We previously devised and developed a novel concept microarray with manageable volumes (MMV) using a soft gel. It demonstrated the great potential of the MMV technology with the examples of 1024-parallel-cell culture and PCR experiments. However, its full potential failed to be expressed, owing to the nature of the material used for the MMV chip. RESULTS: In the present study, by developing plastic-based MMVs and associated technologies, we introduced novel technologies such as C2D2P (in which the cells in each well are converted from DNA to protein in 1024-parallel), NGS-non-dependent microbiome analysis, and other powerful applications. CONCLUSIONS: The reborn MMV-microarray technology has proven to be highly efficient and cost-effective (with approximately 100-fold cost reduction) and enables us to realize hitherto unattainable technologies.

Role of Yoga in Cardiovascular Diseases.

Cardiovascular diseases are a collection of conditions that affect the blood vessels and the heart. These conditions include cardiac rehabilitation, hypertension, cardiac failure, rheumatic heart disease, peripheral artery disease, and coronary heart disease. Poor nutrition, alcohol, lack of exercise, smoking, etc are the main behavioral risk factors for heart disease. This study delivers a methodical review of yoga's role in the management and inhibition of cardiovascular diseases and their associated risk factors. This review suggests that proper maintenance of fitness and stress employing yoga effectively lowers cardiovascular disease. In this review, various asanas, and pranayama like Marjaryasana, Kapalabhati, Halasana, etc have been discussed. Also, their role in the prevention of coronary heart disease (CHD). In addition to this different metabolic syndrome associated with cardiovascular diseases and the relation of yoga with hypertension has been discussed. The review has documented satisfactory proof and concludes that yogic exercise enhances cardiovascular health and reduces associated risk factors.

Engineering Considerations on Large-Scale Cultured Meat Production.

In recent decades, cultured meat has received considerable interest as a sustainable alternative to traditional meat products, showing promise for addressing the inherent problems associated with conventional meat production. However, current limitations on the scalability of production and extremely high production costs have prevented their widespread adoption. Therefore, it is important to develop novel engineering strategies to overcome the current limitations in large-scale cultured meat production. Such engineering considerations have the potential for advancements in cultured meat production by providing innovative and effective solutions to the prevailing challenges. In this review, we discuss how engineering strategies have been utilized to advance cultured meat technology by categorizing the production processes of cultured meat into three distinct steps: (1) cell preparation; (2) cultured meat fabrication; and (3) cultured meat maturation. For each step, we provide a comprehensive discussion of the recent progress and its implications. In particular, we focused on the engineering considerations involved in each step of cultured meat production, with specific emphasis on large-scale production.

Decryption of sequence, structure, and functional features of SINE repeat elements in SINEUP non-coding RNA-mediated post-transcriptional gene regulation.

RNA structure folding largely influences RNA regulation by providing flexibility and functional diversity. In silico and in vitro analyses are limited in their ability to capture the intricate relationships between dynamic RNA structure and RNA functional diversity present in the cell. Here, we investigate sequence, structure and functional features of mouse and human SINE-transcribed retrotransposons embedded in SINEUPs long non-coding RNAs, which positively regulate target gene expression post-transcriptionally. In-cell secondary structure probing reveals that functional SINEs-derived RNAs contain conserved short structure motifs essential for SINEUP-induced translation enhancement. We show that SINE RNA structure dynamically changes between the nucleus and cytoplasm and is associated with compartment-specific binding to RBP and related functions. Moreover, RNA-RNA interaction analysis shows that the SINE-derived RNAs interact directly with ribosomal RNAs, suggesting a mechanism of translation regulation. We further predict the architecture of 18 SINE RNAs in three dimensions guided by experimental secondary structure data. Overall, we demonstrate that the conservation of short key features involved in interactions with RBPs and ribosomal RNA drives the convergent function of evolutionarily distant SINE-transcribed RNAs.

The Unusual Adverse Effects of Antituberculosis Therapy in Kidney Patients.

BACKGROUND: Chronic kidney disease (CKD) patients are at a high risk of tuberculosis (TB), with a relative risk of developing active TB of 10%-25%. Similarly, glomerular disease increases the risk of TB due to diminished glomerular filtration rate, proteinuria, and immunosuppression use. Further, the first-line anti-TB drugs are associated with acute kidney injury (AKI) even in patients with normal kidney functions. METHODS: We retrospectively identified 10 patients hospitalized with unusual adverse effects of antituberculosis therapy (ATT) from 2013 to 2022. RESULTS: We found three cases of AKI caused by rifampicin: acute interstitial nephritis, crescentic glomerulonephritis, and heme pigment-induced acute tubular necrosis. We observed rifampicin-induced accelerated hypertension and thrombocytopenia in two patients on maintenance hemodialysis. Isoniazid caused pancreatitis and cerebellitis in two CKD patients, respectively. In a CKD patient, we detected acute gout secondary to pyrazinamide-induced reduced uric acid excretion. We also observed cases of drug rash with eosinophilia and systemic symptoms and hypercalcemia due to immune reconstitution inflammatory syndrome in patients with glomerular disease on ATT. Immediate discontinuation of the offending drug, along with specific and supportive management, led to a recovery in all cases. CONCLUSION: The adverse effects of ATT may be unusually severe and varied in kidney patients due to decreased renal elimination. Early recognition of these adverse effects and timely discontinuation of the offending drug is essential to limit morbidity and mortality.