Valproic Acid-Induced Upregulation of Multidrug Efflux Transporter ABCG2/BCRP via PPARα-Dependent Mechanism in Human Brain Endothelial Cells.

Despite the progress made in the development of new antiepileptic drugs (AEDs), poor response to them is a rising concern in epilepsy treatment. Of several hypotheses explaining AED treatment failure, the most promising theory is the overexpression of multidrug transporters belonging to ATP-binding cassette (ABC) transporter family at blood-brain barrier. Previous data show that AEDs themselves can induce these transporters, in turn affecting their own brain bioavailability. Presently, this induction and the underlying regulatory mechanism involved at human blood-brain barrier is not well elucidated. Herein, we sought to explore the effect of most prescribed first- and second-line AEDs on multidrug transporters in human cerebral microvascular endothelial cells, hCMEC/D3. Our work demonstrated that exposure of these cells to valproic acid (VPA) induced mRNA, protein, and functional activity of breast cancer resistance protein (BCRP/ABCG2). On examining the substrate interaction status of AEDs with BCRP, VPA, phenytoin, and lamotrigine were found to be potential BCRP substrates. Furthermore, we observed that siRNA-mediated knockdown of peroxisome proliferator-activated receptor alpha (PPARα) or use of PPARα antagonist, resulted in attenuation of VPA-induced BCRP expression and transporter activity. VPA was found to increase PPARα expression and trigger its translocation from cytoplasm to nucleus. Findings from chromatin immunoprecipitation and luciferase assays showed that VPA enhances the binding of PPARα to its response element in the ABCG2 promoter, resulting in elevated ABCG2 transcriptional activity. Taken together, these in vitro findings highlight PPARα as the potential molecular target to prevent VPA-mediated BCRP induction, which may have important implications in VPA pharmacoresistance. SIGNIFICANCE STATEMENT: Induction of multidrug transporters at blood-brain barrier can largely affect the bioavailability of the substrate antiepileptic drugs in the brains of patients with epilepsy, thus affecting their therapeutic efficacy. The present study reports a mechanistic pathway of breast cancer resistance protein (BCRP/ABCG2) upregulation by valproic acid in human brain endothelial cells via peroxisome proliferator-activated receptor alpha involvement, thereby providing a potential strategy to prevent valproic acid pharmacoresistance in epilepsy.

Adhatoda vasica rescues the hypoxia-dependent severe asthma symptoms and mitochondrial dysfunction.

Severe asthma is a chronic airway disease that exhibits poor response to conventional asthma therapies. Growing evidence suggests that elevated hypoxia increases the severity of asthmatic inflammation among patients and in model systems. In this study, we elucidate the therapeutic effects and mechanistic basis of Adhatoda vasica (AV) aqueous extract on mouse models of acute allergic as well as severe asthma subtypes at physiological, histopathological, and molecular levels. Oral administration of AV extract attenuates the increased airway resistance and inflammation in acute allergic asthmatic mice and alleviates the molecular signatures of steroid (dexamethasone) resistance like IL-17A, KC (murine IL-8 homologue), and HIF-1α (hypoxia-inducible factor-1α) in severe asthmatic mice. AV inhibits HIF-1α levels through restoration of expression of its negative regulator-PHD2 (prolyl hydroxylase domain-2). Alleviation of hypoxic response mediated by AV is further confirmed in the acute and severe asthma model. AV reverses cellular hypoxia-induced mitochondrial dysfunction in human bronchial epithelial cells-evident from bioenergetic profiles and morphological analysis of mitochondria. In silico docking of AV constituents reveal higher negative binding affinity for C and O-glycosides for HIF-1α, IL-6, Janus kinase 1/3, TNF-α, and TGF-ß-key players of hypoxia inflammation. This study for the first time provides a molecular basis of action and effect of AV whole extract that is widely used in Ayurveda practice for diverse respiratory ailments. Further, through its effect on hypoxia-induced mitochondrial dysfunction, the study highlights its potential to treat severe steroid-resistant asthma.

Beclin 1 controls pigmentation by changing the nuclear localization of melanogenic factor MITF.

The primary contributor for the determination of skin color is melanin, a pigment that is produced in specialized cells called melanocytes. At cellular level, melanin synthesis occurs through several enzymes like tyrosinase (TYR) and tyrosinase related proteins and the expression of these proteins are regulated transcriptionally by microphthalmia associated transcription factor (MITF). Melanin pigmentation is a complex process finely regulated by different transcription factors, structural proteins and enzymes. In recent times, several autophagic genes have been implicated in the regulation of pigmentation. Though previous report observed a visible loss of coat-color in heterozygous Beclin 1 mice, the role of this protein in pigmentation is yet to study in details. In this present work we intend to study the role of Beclin 1, a central autophagic factor, in pigmentation. Using human melanoma cells and primary melanocytes, we showed that Beclin 1 downregulation significantly decreased the melanin content, tyrosinase activity and the expression of TYR and tyrosinase related protein 1 (TYRP1). These effects were recapitulated in a Beclin 1 knockdown in vivo model of zebrafish. Most importantly, re-expression of Beclin 1 rescued the pigmentation-associated defects both in cellular and in organismal level indicating the specificity. Surprisingly, Beclin 1 knockdown cells did not show significant changes in MITF expression but the nuclear localization of MITF was altered. Together, these data suggest that indeed Beclin 1 is associated with melanogenesis and this effect is more likely exerted through the subcellular distribution rather than the change in expression of MITF.

Encephalopathy due to defective mitochondrial and peroxisomal fission 2 caused by a novel MFF gene mutation in a young child.

Encephalopathy due to defective mitochondrial and peroxisomal fission 2 caused by mitochondrial fission factor (MFF) gene mutation is a rare neurogenetic disorder. Pathogenic MFF mutations have been described in three reports in literature so far. We report a young child of Indian descent who presented to us with global developmental followed by regression of acquired milestones, spasticity, visual and auditory impairment, and was found to harbor a novel pathogenic homozygous MFF truncating variant c.433C>T; p.Arg145Ter. Cellular imaging of patient lymphoblastoid cell line had shown abnormal shapes of mitochondria due to fission defects. The patient has been started on mitochondrial cocktail with some improvement.

Optimizing Anesthesia for Extensive Extraoral Fungating Lesions: Strategies and Considerations.

A large extraoral fungus, frequently seen in late head and neck cancers, poses serious difficulties for the management of anesthesia and surgery. Essential factors include preoperative optimization, airway assessment, intraoperative monitoring, and postoperative care. Risk mitigation and outcome optimization strategies are discussed, including appropriate airway management and hemodynamic monitoring. Ideal patient outcomes in situations of extensive extraoral fungation can be attained by a complete plan that integrates surgical expertise and anesthetic care. This case discusses the successful anesthetic management of a 55-year-old man undergoing composite resection with segmental mandibulectomy, appropriate neck dissection, free fibular flap, and scalp flap for squamous cell carcinoma of the lower labial mucosa with significant extraoral fungation.

A Case Presentation of Well-Differentiated Hepatocellular Carcinoma With No Sign of Liver Disease.

The type of liver cancer that occurs most frequently is hepatocellular carcinoma (HCC). The majority of cases of HCC are secondary to alcoholic cirrhosis or viral hepatitis. The presence of malignant cells with modest nuclear atypia that resemble normal hepatocytes and the lack of bare nuclei in the smears, which shows the neoplastic hepatocytes' capacity, are characteristics of a well-differentiated HCC plasma membrane to tolerate smearing. We present the case of an 83-year-old male patient with a well-differentiated HCC, who had no etiological factors and no signs of alcohol cirrhotic liver, or any symptoms of liver disease which are the main causes of the HCC.

Unusual Presentation of a Giant Cell Tumor of the Bone: A Case Report.

Bone giant cell tumors (GCTs) are rare, non-cancerous tumors that mostly affect the meta-epiphyseal region of long bones in the legs and arms. We are reporting a case of GCT of bone of a 14-year-old male; it usually occurs in the age group of 20-40 years. The presence of multinucleated giant cells and stromal cells in the proximal diaphysis of the left tibia serves as a distinguishing characteristic. The majority of GCTs are benign; they have the potential to induce bone loss and can be locally aggressive. Treatment options often include surgery, and in some cases, medications like denosumab may be used to help shrink the tumor or manage recurrent cases.

Role of Cathepsin B Expression in Oral Squamous Cell Carcinoma: A Comprehensive Review.

This comprehensive review delves into the intricate landscape of oral squamous cell carcinoma (OSCC) by examining the role of cathepsin B expression in its pathogenesis. OSCC, a prevalent and clinically significant oral malignancy, poses a considerable global health burden, necessitating a thorough exploration of its underlying molecular mechanisms. Cathepsin B, a lysosomal cysteine protease, emerges as a critical player in OSCC, influencing tumour initiation, invasion, and metastasis. The review begins with a brief overview of OSCC, emphasizing its epidemiological and clinical features, followed by exploring the significance of studying cathepsin B expression in this context. In the manuscript, the structure and function of cathepsin B are elucidated, providing a foundation for understanding its aberrant expression in OSCC. Clinical studies revealing correlations with tumour grade and stage, along with prognostic significance, are scrutinized, offering insights into the potential diagnostic and prognostic utility of cathepsin B. The biological functions of cathepsin B in OSCC, including its impact on tumour invasion and modulation of apoptosis, are comprehensively discussed. The Therapeutic Implications section explores targeting cathepsin B as a potential strategy, emphasizing the need for future research to overcome associated challenges. In the Conclusion section, the review synthesizes key findings, delineates implications for future research, and highlights the potential impact of cathepsin B on OSCC diagnosis and treatment, contributing to the ongoing efforts to advance our understanding of this complex malignancy, which is associated with a high mortality rate and improve clinical outcomes.

Beyond the Norm: Navigating a Unique Papillary Carcinoma Journey in Breast Cancer.

The presence of papillary structures inside the tumor is a unique and uncommon characteristic of breast cancer, and it is known as papillary carcinoma. In contrast to other forms of breast cancer, this variant usually manifests as a well-defined mass in imaging investigations and is frequently linked to a good prognosis. We present a case of a 72-year-old female with papillary carcinoma of the breast identified after presenting with a palpable breast lump. Following a left simple mastectomy and adjuvant treatment, the presence of papillary structures inside the tumor was verified by a histopathological study. Understanding the clinical and pathological characteristics of breast papillary carcinoma is crucial for precise diagnosis and suitable therapy strategizing. More research is required to further understand the molecular traits and best practices for treating this uncommon subtype of breast cancer.

Stealthy Encroachment: Unraveling an Encounter With Renal Myelolipoma.

Myelolipoma of the kidney is an exceedingly unusual benign tumor of hematological components mixed with mature adipose tissue. We present a case of a 59-year-old male who presented with left flank pain and was found to have an atrophic left kidney on imaging studies. A computed tomography (CT) scan revealed a small and shrunken left kidney with an extrarenal pelvis. A diethylenetriamine pentaacetate (DTPA) scan results showed a total glomerular filtration rate (GFR) of 45.6 ml/min with a non-functional left kidney. The patient underwent a left nephrectomy, and a histopathological examination confirmed the diagnosis and highlighted the distinctive morphological features of this rare entity. Postoperatively, the patient experienced a complete resolution of symptoms. This case underscores the importance of considering myelolipoma in the differential diagnosis of renal masses and highlights the successful management of symptomatic cases through surgical intervention. Awareness of this rare tumor is crucial for accurate diagnosis and appropriate management. Further studies are needed to elucidate the natural history and optimal treatment strategies for renal myelolipomas.

From Pixels to Prognosis: A Narrative Review on Artificial Intelligence's Pioneering Role in Colorectal Carcinoma Histopathology.

Colorectal carcinoma, a prevalent and deadly malignancy, necessitates precise histopathological assessment for effective diagnosis and prognosis. Artificial intelligence (AI) emerges as a transformative force in this realm, offering innovative solutions to enhance traditional histopathological methods. This narrative review explores AI's pioneering role in colorectal carcinoma histopathology, encompassing its evolution, techniques, and advancements. AI algorithms, notably machine learning and deep learning, have revolutionized image analysis, facilitating accurate diagnosis and prognosis prediction. Furthermore, AI-driven histopathological analysis unveils potential biomarkers and therapeutic targets, heralding personalized treatment approaches. Despite its promise, challenges persist, including data quality, interpretability, and integration. Collaborative efforts among researchers, clinicians, and AI developers are imperative to surmount these hurdles and realize AI's full potential in colorectal carcinoma care. This review underscores AI's transformative impact and implications for future oncology research, clinical practice, and interdisciplinary collaboration.

Cardiovascular aging: the mitochondrial influence.

Age-associated cardiovascular disease is becoming progressively prevalent due to the increased lifespan of the population. However, the fundamental mechanisms underlying the aging process and the corresponding decline in tissue functions are still poorly understood. The heart has a very high energy demand and the cellular energy needed to sustain contraction is primarily generated by mitochondrial oxidative phosphorylation. Mitochondria are also involved in supporting various metabolic processes, as well as activation of the innate immune response and cell death pathways. Given the central role of mitochondria in energy metabolism and cell survival, the heart is highly susceptible to the effects of mitochondrial dysfunction. These key organelles have been implicated as underlying drivers of cardiac aging. Here, we review the evidence demonstrating the mitochondrial contribution to the cardiac aging process and disease susceptibility. We also discuss the potential mechanisms responsible for the age-related decline in mitochondrial function.

Obesity impairs cardiolipin-dependent mitophagy and therapeutic intercellular mitochondrial transfer ability of mesenchymal stem cells.

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.

Mitochondria are secreted in extracellular vesicles when lysosomal function is impaired.

Mitochondrial quality control is critical for cardiac homeostasis as these organelles are responsible for generating most of the energy needed to sustain contraction. Dysfunctional mitochondria are normally degraded via intracellular degradation pathways that converge on the lysosome. Here, we identified an alternative mechanism to eliminate mitochondria when lysosomal function is compromised. We show that lysosomal inhibition leads to increased secretion of mitochondria in large extracellular vesicles (EVs). The EVs are produced in multivesicular bodies, and their release is independent of autophagy. Deletion of the small GTPase Rab7 in cells or adult mouse heart leads to increased secretion of EVs containing ubiquitinated cargos, including intact mitochondria. The secreted EVs are captured by macrophages without activating inflammation. Hearts from aged mice or Danon disease patients have increased levels of secreted EVs containing mitochondria indicating activation of vesicular release during cardiac pathophysiology. Overall, these findings establish that mitochondria are eliminated in large EVs through the endosomal pathway when lysosomal degradation is inhibited.

NSP4 and ORF9b of SARS-CoV-2 Induce Pro-Inflammatory Mitochondrial DNA Release in Inner Membrane-Derived Vesicles.

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.

Cellular and mitochondrial calcium communication in obstructive lung disorders.

Calcium (Ca2+) signalling is well known to dictate cellular functioning and fate. In recent years, the accumulation of Ca2+ in the mitochondria has emerged as an important factor in Chronic Respiratory Diseases (CRD) such as Asthma and Chronic Obstructive Pulmonary Disease (COPD). Various reports underline an aberrant increase in the intracellular Ca2+, leading to mitochondrial ROS generation, and further activation of the apoptotic pathway in these diseases. Mitochondria contribute to Ca2+ buffering which in turn regulates mitochondrial metabolism and ATP production. Disruption of this Ca2+ balance leads to impaired cellular processes like apoptosis or necrosis and thus contributes to the pathophysiology of airway diseases. This review highlights the key role of cytoplasmic and mitochondrial Ca2+ signalling in regulating CRD, such as asthma and COPD. A better understanding of the dysregulation of mitochondrial Ca2+ homeostasis in these diseases could provide cues for the development of advanced therapeutic interventions in these diseases.

Cardiolipin-mediated PPARγ S112 phosphorylation impairs IL-10 production and inflammation resolution during bacterial pneumonia.

Bacterial pneumonia is a global healthcare burden, and unwarranted inflammation is suggested as an important cause of mortality. Optimum levels of the anti-inflammatory cytokine IL-10 are essential to reduce inflammation and improve survival in pneumonia. Elevated levels of the mitochondrial-DAMP cardiolipin (CL), reported in tracheal aspirates of pneumonia patients, have been shown to block IL-10 production from lung MDSCs. Although CL-mediated K107 SUMOylation of PPARγ has been suggested to impair this IL-10 production, the mechanism remains elusive. We identify PIAS2 to be the specific E3-SUMOligase responsible for this SUMOylation. Moreover, we identify a concomitant CL-mediated PPARγ S112 phosphorylation, mediated by JNK-MAPK, to be essential for PIAS2 recruitment. Furthermore, using a clinically tested peptide inhibitor targeting JNK-MAPK, we blocked these post-translational modifications (PTMs) of PPARγ and rescued IL-10 expression, improving survival in murine pneumonia models. Thus, we explore the mechanism of mito-DAMP-mediated impaired lung inflammation resolution and propose a therapeutic strategy targeting PPARγ PTMs.

Generation of induced pluripotent stem cell line (IGIBi007-A) from a patient with a novel acromesomelic dysplasia, PRKG2 type (AMDP).

Biallelic PRKG2 (Protein Kinase, cGMP dependent Type-2) mutations cause a novel acromesomelic dysplasia PRKG2 type. We report generation of induced pluripotent stem cell line from lymphoblastoid cell lines of the patient carrying the reported frameshift mutation (p.Asn164Lysfs*2). The derived iPSC line exhibits all the features of pluripotency, free of major genetic alterations due to reprogramming process and has the capability to differentiate into three germ layers. This iPSC cell line may provide an opportunity to investigate the effect of PRKG2 mutations upon FGF (fibroblast-growth-factor) induced MAPK signalling involved in chondrocyte proliferation in-vitro and may aid in possible therapeutic screening of novel biomolecules.

Probiotic Bacteria in Wound Healing; An In-Vivo Study.

BACKGROUND: Probiotics are food supplements that benefit the host by improving its intestinal microbial balance. Probiotics are used as diet supplements to prevent diarrhoea and improve lactose tolerance. OBJECTIVES: The present study deals with the isolation of a potent probiotic strain capable of inducing healing properties in rat model. MATERIALS AND METHODS: Probiotic VITSAMJ1 was isolated from goat milk using MRS media. The antimicrobial assay was carried out against S. aureus (MTCC 3160) and the wound healing properties were assessed on female Wistar rats. A 1.5 cm2 subcutaneous wound was induced in the rats, and a probiotic gel formulation was tropically applied onto the wounds. Tissue biopsy was carried out after days 1, 3, 5, 7, 9, and 11. Total leucocyte count and Histopathological analysis were performed after each interval. RESULTS: VITSAMJ1 can be effectively used for wound healing. CONCLUSION: VITSAMJ1 can be effectively used for wound healing.