Extracellular vesicle-mediated delivery of anti-miR-106b inhibits morphine-induced primary ciliogenesis in the brain.

Repeated use of opioids such as morphine causes changes in the shape and signal transduction pathways of various brain cells, including astrocytes and neurons, resulting in alterations in brain functioning and ultimately leading to opioid use disorder. We previously demonstrated that extracellular vesicle (EV)-induced primary ciliogenesis contributes to the development of morphine tolerance. Herein, we aimed to investigate the underlying mechanisms and potential EV-mediated therapeutic approach to inhibit morphine-mediated primary ciliogenesis. We demonstrated that miRNA cargo in morphine-stimulated-astrocyte-derived EVs (morphine-ADEVs) mediated morphine-induced primary ciliogenesis in astrocytes. CEP97 is a target of miR-106b and is a negative regulator of primary ciliogenesis. Intranasal delivery of ADEVs loaded with anti-miR-106b decreased the expression of miR-106b in astrocytes, inhibited primary ciliogenesis, and prevented the development of tolerance in morphine-administered mice. Furthermore, we confirmed primary ciliogenesis in the astrocytes of opioid abusers. miR-106b-5p in morphine-ADEVs induces primary ciliogenesis via targeting CEP97. Intranasal delivery of ADEVs loaded with anti-miR-106b ameliorates morphine-mediated primary ciliogenesis and prevents morphine tolerance. Our findings bring new insights into the mechanisms underlying primary cilium-mediated morphine tolerance and pave the way for developing ADEV-mediated small RNA delivery strategies for preventing substance use disorders.

Changes in Plasma Metabolic Signature upon Acute and Chronic Morphine Administration in Morphine-Tolerant Mice.

Morphine administration causes system-level metabolic changes. Here, we show that morphine-tolerant mice exhibited distinct plasma metabolic signatures upon acute and chronic administration. We utilized a mouse model of morphine tolerance by exposing mice to increasing doses of the drug over 4 days. We collected plasma samples from mice undergoing acute or chronic morphine or saline injections and analyzed them using targeted GC-MS-based metabolomics to profile approximately 80 metabolites involved in the central carbon, amino acid, nucleotide, and lipid metabolism. Our findings reveal distinct alterations in plasma metabolite concentrations in response to acute or chronic morphine intake, and these changes were linked to the development of tolerance to morphine's analgesic effects. We identified several metabolites that had been differentially affected by acute versus chronic morphine use, suggesting that metabolic changes may be mitigated by prolonged exposure to the drug. Morphine-tolerant mice showed a restoration of amino acid and glycolytic metabolites. Additionally, we conducted reconstructed metabolic network analysis on the first 30 VIP-ranked metabolites from the PLSDA of the saline, acute, and morphine-tolerant mice groups, which uncovered four interaction networks involving the amino acid metabolism, the TCA cycle, the glutamine-phenylalanine-tyrosine pathway, and glycolysis. These pathways were responsible for the metabolic differences observed following distinct morphine administration regimens. Overall, this study provides a valuable resource for future investigations into the role of metabolites in morphine-induced analgesia and associated effects following acute or chronic use in mice.

Deciphering sperm functions using biological networks.

The global human population is exponentially increasing, which requires the production of quality food through efficient reproduction as well as sustainable production of livestock. Lack of knowledge and technology for assessing semen quality and predicting bull fertility is hindering advances in animal science and food animal production and causing millions of dollars of economic losses annually. The intent of this systemic review is to summarize methods from computational biology for analysis of gene, metabolite, and protein networks to identify potential markers that can be applied to improve livestock reproduction, with a focus on bull fertility. We provide examples of available gene, metabolic, and protein networks and computational biology methods to show how the interactions between genes, proteins, and metabolites together drive the complex process of spermatogenesis and regulate fertility in animals. We demonstrate the use of the National Center for Biotechnology Information (NCBI) and Ensembl for finding gene sequences, and then use them to create and understand gene, protein and metabolite networks for sperm associated factors to elucidate global cellular processes in sperm. This study highlights the value of mapping complex biological pathways among livestock and potential for conducting studies on promoting livestock improvement for global food security.

Clinicopathological Profile of Non-small Cell Lung Cancer and the Changing Trends in Its Histopathology: Experience From a Tertiary Care Cancer Center in Kashmir, India.

Background The overall frequency and incidence of different cancers across the globe, including lung cancer, are marked by ethnic and geographical variations. Lung cancer is the most commonly diagnosed cancer worldwide that inflicts most of the cancer deaths. Non-small cell lung cancer (NSCLC) constitutes most lung cancer cases. The aim of this study was to find the frequency and clinicopathological characteristics of NSCLC in high incidence zone of Kashmir, an ethnically and geographically distinct area in Northern India. Material and methods The study was conducted to evaluate the clinicopathological profile of NSCLC at a tertiary care cancer center, Sher-I-Kashmir Institute of Medical Sciences (SKIMS). The patients and case records were analyzed for clinical presentation and demographic features, smoking status, radiological features, histopathological type, and stage of their disease at presentation. Results The study included 1557 NSCLC patients registered over a period of seven years, i.e., 2008-2014. Most of the patients belonged to rural areas (70%). The median age of the whole cohort was 58.0 years (ranges 22-95 years), and the male-to-female ratio was 3.7:1 (male = 1231 and female = 326). Smokers comprised 77.39% of cases, and Hookah was the most common form of smoking (65.06%). The ratio of squamous cell carcinoma and adenocarcinoma was 3.7:1 (67.5% vs. 24.9%). Stage III and IV disease accounted for 93% of cases (30.6% and 62.7%, respectively). Squamous cell carcinoma histopathology was dominant in smokers (74.3%) compared to adenocarcinoma (19%), while squamous cell carcinoma and adenocarcinoma histopathology ran parallel in nonsmokers (45.1% and 44%, respectively). Most of the patients had an Eastern Cooperative Oncology Group (ECOG) performance status between 1 and 2 (79%).  Conclusion We conclude Kashmir region is a very high-risk area for lung cancer, with NSCLC showing a high incidence. Most of our patients present in advanced stages, and the frequency of adenocarcinoma is showing an increasing trend over the years from 2008-2014.

Diagnostic implication of a circulating serum-based three-microRNA signature in hepatocellular carcinoma.

Owing to the diagnostic dilemma, the prognosis of hepatocellular carcinoma (HCC) remains impoverished, contributing to the globally high mortality rate. Currently, HCC diagnosis depends on the combination of imaging modalities and the measurement of serum alpha-fetoprotein (AFP) levels. Nevertheless, these conventional modalities exhibit poor performance in detecting HCC at early stages. Thus, there is a pressing need to identify novel circulating biomarkers to promote diagnostic accuracy and surveillance. Circulating miRNAs are emerging as promising diagnostic tools in screening various cancers, including HCC. However, because of heterogenous and, at times, contradictory reports, the universality of miRNAs in clinical settings remains elusive. Consequently, we proposed to explore the diagnostic potential of ten miRNAs selected on a candidate-based approach in HCC diagnosis. The expression of ten candidate miRNAs (Let-7a, miR-15a, miR-26a, miR-124, miR-126, miR-155, miR-219, miR-221, miR-222, and miR-340) was investigated in serum and tissue of 66 subjects, including 33 HCC patients and 33 healthy controls (HC), by rt-PCR. Receiver operating characteristic curve (ROC) analysis was used to determine the diagnostic accuracy of the prospective serum miRNA panel. To anticipate the potential biological roles of a three-miRNA signature, the target genes were evaluated using the Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway. The serum and tissue expression of miRNAs (Let-7a, miR-26a, miR-124, miR-155, miR-221, miR-222, and miR-340) were differentially expressed in HCC patients (p < 0.05). The ROC analysis revealed promising diagnostic performance of Let-7a (AUC = 0.801), miR-221 (AUC = 0.786), and miR-2 (AUC = 0.758) in discriminating HCC from HC. Furthermore, in a logistic regression equation, we identified a three-miRNA panel (Let-7a, miR-221, and miR-222; AUC = 0.932) with improved diagnostic efficiency in differentiating HCC from HC. Remarkably, the combination of AFP and a three-miRNA panel offered a higher accuracy of HCC diagnosis (AUC = 0.961) than AFP alone. The functional enrichment analysis demonstrated that target genes may contribute to pathways associated with HCC and cell-cycle regulation, indicating possible crosstalk of miRNAs with HCC development. To conclude, the combined classifier of a three-miRNA panel and AFP could be indispensable circulating biomarkers for HCC diagnosis. Furthermore, targeting predicted genes may provide new therapeutic clues for the treatment of aggressive HCC.

Percutaneous Transluminal Angioplasty of Dysfunctional Hemodialysis Vascular Access: Can Careful Selection of Patients Improve the Outcomes?

Introduction: Our study aimed to evaluate the role of endovascular intervention in salvaging hemodialysis access in patients of end-stage renal disease with specific attention to features that may predict a poor outcome. We also evaluated the role of ultrasonography (USG) in the management of these patients. Methods: Forty-two patients with dysfunctional hemodialysis arteriovenous fistulas (AVF) were taken up for percutaneous transluminal angioplasty (PTA) with or without stent placement. All patients underwent a pre- and postprocedural USG Doppler to assess parameters such as mean flow, mean peak systolic velocity, and vessel diameter. Technical and clinical success rates were calculated, and characteristics causing increased failure rates (long-segment and multisite stenosis and diabetes) were noted. Results: The most common sites of stenosis were the anastomotic and perianastomotic sites (n = 27, 63%) on the venous side followed by distal venous drainage site (23%) and central venous stenosis (14%). The technical and clinical success rates were 98% and 92%, respectively. Three- and 6-month patency rates were 83% and 71%, respectively. Common characteristics in patients with failure (primary or secondary) were diabetes, increased age, increased length of stenosis (>2cm) and multisite stenosis. USG Doppler parameters showed a significant improvement post-PTA (P < 0.001) indicating clinical success. No major complication was noted in our study. Conclusion: PTA is successful for dysfunctional hemodialysis access. Careful selection of patients can improve the success rates and decrease economic burden in a resource-constrained country like ours. USG Doppler is essential in the assessment of iatrogenic hemodialysis AVFs.

Extracellular Vesicle-Mediated Delivery of Ultrasmall Superparamagnetic Iron Oxide Nanoparticles to Mice Brain.

Extracellular vesicles (EVs) are small lipid membrane-bound vesicles that can pass the blood-brain barrier. Therefore, EVs could be used for the delivery of therapeutics to the brain. Herein, we investigated the biodistribution of intranasal perfusion of ultrasmall superparamagnetic iron oxide (USPIO)-labeled astrocyte-derived EVs (ADEVs) in mice. We used Western blotting, transmission electron microscopy (TEM), and nanoparticle uptake assay to characterize ADEVs. In addition, intranasal perfusion coupled with magnetic resonance imaging (MRI) was employed to determine the distribution of USPIO-labeled ADEVs in mice. Our results showed the uptake of USPIO by mouse astrocytes and ADEVs. In addition, we confirmed the biodistribution of ADEVs in the brain and other internal organs, including the kidneys, liver, and spleen. Our results suggest that USPIO did not affect mouse astrocyte cell survivability and EV release. Therefore, intranasal delivery of therapeutic loaded EVs could be used for the treatment of various brain disorders.

Primary cilia and ciliary signaling pathways in aging and age-related brain disorders.

Brain disorders are characterized by the progressive loss of structure and function of the brain as a consequence of progressive degeneration and/or death of nerve cells. Aging is a major risk factor for brain disorders such as Alzheimer's disease (AD), Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), and stroke. Various cellular and molecular events have been shown to play a role in the progress of neurodegenerative diseases. Emerging studies suggest that primary cilia could be a key regulator in brain diseases. The primary cilium is a singular cellular organelle expressed on the surface of many cell types, such as astrocytes and neurons in the mature brain. Primary cilia detect extracellular cues, such as Sonic Hedgehog (SHH) protein, and transduce these signals into cells to regulate various signaling pathways. Abnormalities in ciliary length and frequency (ratio of ciliated cells) have been implicated in various human diseases, including brain disorders. This review summarizes current findings and thoughts on the role of primary cilia and ciliary signaling pathways in aging and age-related brain disorders.

Role of Autophagy in HIV-1 and Drug Abuse-Mediated Neuroinflammaging.

Chronic low-grade inflammation remains an essential feature of HIV-1 infection under combined antiretroviral therapy (cART) and contributes to the accelerated cognitive defects and aging in HIV-1 infected populations, indicating cART limitations in suppressing viremia. Interestingly, ~50% of the HIV-1 infected population on cART that develops cognitive defects is complicated by drug abuse, involving the activation of cells in the central nervous system (CNS) and neurotoxin release, altogether leading to neuroinflammation. Neuroinflammation is the hallmark feature of many neurodegenerative disorders, including HIV-1-associated neurocognitive disorders (HAND). Impaired autophagy has been identified as one of the underlying mechanisms of HAND in treated HIV-1-infected people that also abuse drugs. Several lines of evidence suggest that autophagy regulates CNS cells' responses and maintains cellular hemostasis. The impairment of autophagy is associated with low-grade chronic inflammation and immune senescence, a known characteristic of pathological aging. Therefore, autophagy impairment due to CNS cells, such as neurons, microglia, astrocytes, and pericytes exposure to HIV-1/HIV-1 proteins, cART, and drug abuse could have combined toxicity, resulting in increased neuroinflammation, which ultimately leads to accelerated aging, referred to as neuroinflammaging. In this review, we focus on the potential role of autophagy in the mechanism of neuroinflammaging in the context of HIV-1 and drug abuse.

Astrocyte-Derived Extracellular Vesicle-Mediated Activation of Primary Ciliary Signaling Contributes to the Development of Morphine Tolerance.

BACKGROUND: Morphine is used extensively in the clinical setting owing to its beneficial effects, such as pain relief; its therapeutic utility is limited because the prolonged use of morphine often results in tolerance and addiction. Astrocytes in the brain are a direct target of morphine action and play an essential role in the development of morphine tolerance. Primary cilia and the cilia-mediated sonic hedgehog (SHH) signaling pathways have been shown to play a role in drug resistance and morphine tolerance, respectively. Extracellular vesicles (EVs) play important roles as cargo-carrying vesicles mediating communication among cells and tissues. METHODS: C57BL/6N mice were administered morphine for 8 days to develop tolerance, which was determined using the tail-flick and hot plate assays. EVs were separated from astrocyte-conditioned media using either size exclusion chromatography or ultracentrifugation approaches, followed by characterization of EVs using nanoparticle tracking analysis for EV size distribution and number, Western blotting for EV markers, and electron microscopy for EV morphology. Astrocytes were treated with EVs for 24 hours, followed by assessing primary cilia by fluorescent immunostaining for primary cilia markers (ARL13B and acetylated tubulin). RESULTS: Morphine-tolerant mice exhibited an increase in primary cilia length and percentage of ciliated astrocytes. The levels of SHH protein were upregulated in morphine-stimulated astrocyte-derived EVs. SHH on morphine-stimulated astrocyte-derived EVs activated SHH signaling in astrocytes through primary cilia. Our in vivo study demonstrated that inhibition of either EV release or primary cilia prevents morphine tolerance in mice. CONCLUSIONS: EV-mediated primary ciliogenesis contributes to the development of morphine tolerance.

Diffusion-Weighted Imaging: An Exciting and Problem-Solving Tool in Patients with Hepatic Metastases.

Background The diffusion-weighted imaging (DWI) is based on the random Brownian motion of water molecules that influences image contrast depending on different pathological conditions. Objective The purpose of this study was to evaluate the efficacy of various magnetic resonance imaging (MRI) sequences including diffusion-weighted and gadobenate-enhanced MRI in the detection and characterization of liver lesions in a patient of known primary malignancy and to compare MRI with contrast-enhanced computed tomography (CECT) and ultrasonography (USG) in the detection of liver metastases. Methods All patients underwent a multiphase MRI. The final diagnosis was established by histopathological examination. Results A total of 43 patients of known primary malignancy were enrolled. MRI gave a provisional diagnosis of liver metastases in 21 patients and benign disease in 22 patients with histopathological correlation revealing two false-negative and one false-positive result. In the detection of lesions, DWI outscored other sequences (92.9 vs. 83.5% in hepatobiliary phase vs. 55.0% in T 2 -weighted sequences) with a statistically significant difference noted only in comparison with T 2 -weighted sequences ( p < 0.001). In 16 patients, MRI added new lesions that were not detected by CECT/USG. The sensitivity and specificity of MRI for detecting metastases were 90.9%/95.2% and 97.9%/96.8% for per-patient and per-lesion basis, respectively. Conclusion Multiphase MRI improved both the detection and characterization of liver metastases. Adding DWI to the routine MR sequences helped in detecting small liver metastases (<10 mm) not detected by other sequences.

The effectiveness of image-guided percutaneous catheter drainage in the management of acute pancreatitis-associated pancreatic collections.

PURPOSE: Acute pancreatitis is commonly complicated by the development of pancreatic collections (PCs). Symptomatic PCs warrant drainage, and the available options include percutaneous, endoscopic, and open surgical approaches. The study aimed to assess the therapeutic effectiveness and safety of image guided percutaneous catheter drainage (PCD) in the management of acute pancreatitis related PCs. MATERIAL AND METHODS: This was a single-centre prospective study covering a 4-year study period. Acute pancreatitisrelated PCs complicated by secondary infection or those producing symptoms due to pressure effect on surrounding structures were enrolled and underwent ultrasound or computed tomography (CT)-guided PCD. The patients were followed to assess the success of PCD (defined as clinical, radiological improvement, and the avoidance of surgery) and any PCD-related complications. RESULTS: The study included 60 patients (60% males) with a mean age of 43.1 ± 21.2 years. PCD recorded a success rate of 80% (16/20) for acute peripancreatic fluid collections (APFC) and pancreatic pseudocysts (PPs), 75% (12/16) for walled-off necrosis (WON), and 50% (12/24) for acute necrotic collections (ANCs). Post-PCD surgery (necrosectomy ± distal pancreatectomy) was needed in 50% of ANC and 25% of WON. Only 20% of APFCs/PPs patients required surgical/endoscopic treatment post-PCD. Minor procedure-related complications were seen in 4 (6.6%) patients. CONCLUSION: PCD is an effective, safe, and minimally invasive therapeutic modality with a good success rate in the management of infected/symptomatic PCs.

Spectrum of chest CT manifestations of coronavirus disease (COVID-19): A pictorial essay.

Coronavirus disease (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is an enveloped single-stranded RNA virus belonging to the family of betacoronaviruses. Chest computed tomography (CT) has helped us in understanding this new disease. Typical CT features of COVID-19 pneumonia are ground-glass opacities (GGO), crazy paving pattern and GGO with superimposed consolidation with a basal, posterior and peripheral lung predilection. Less commonly bronchial wall thickening, bronchial dilatation and pleural thickening are seen. Presence of pleural effusion, pericardial effusion and mediastinal lymphadenopathy is seen in severe cases. Reticulations, fibrous stripes, reverse halo sign and perilobular opacities are seen late (>2 weeks) in the course of illness. We aim to present a pictorial review of CT imaging findings in COVID-19 to illustrate the typical and atypical manifestations of this disease in a bid to familiarize radiologists with the myriad imaging manifestations of this disease.

The spectrum of imaging findings in pulmonary hydatid disease and the additive value of T2-weighted magnetic resonance imaging in its diagnosis.

PURPOSE: To describe the spectrum of imaging findings in pulmonary echinococcosis and to study the additive value of T2-weighted magnetic resonance imaging (MRI) in the characterisation of pulmonary hydatid disease. MATERIAL AND METHODS: This was a descriptive, prospective study conducted for a period of 3 years from December 2016 to November 2019. Patients suspected of having pulmonary echinococcosis (n = 110) on preliminary chest radiography were examined with chest computed tomography (CT). Among them 41 cases were additionally examined with T2-weighted MRI of thorax. Final diagnosis was based on surgery or histopathology. RESULTS: Of the 110 patients enrolled for the study 15 were lost to attrition, and among the final cohort of 95 patients CT correctly diagnosed 68/84 (80.9%) as hydatid cyst, whereas 16/84 (19.1%) received an erroneous alternate diagnosis on CT. Based on the classical findings of hyperintense pulmonary cystic lesion with T2-weighted hypointense rim or detached internal T2-weighted hypointense membrane, a correct diagnosis of hydatid cyst was possible in 30 patients whereas a correct alternate diagnosis was made in 8 cases. T2-weighted MRI was found to have sensitivity of 96.7%, specificity of 80%, positive predictive value (PPV) of 93.7% and negative predictive value (NPV) of 88.9% with an overall diagnostic accuracy of 92.6%. Using the McNemar test, MRI was found to be diagnostically superior to CT (p = 0.019). CONCLUSIONS: Most of the pulmonary hydatid cysts can be diagnosed on CT; however, sometimes the findings may be indeterminate or atypical, leading to a diagnostic dilemma. MRI, owing to its ability to demonstrate hypointense endocyst, can act as a useful adjunct to correctly diagnose hydatid cyst or suggest an alternative diagnosis.

Prolactinoma Outcome After Pregnancy and Lactation: A Cohort Study.

Context: Prolactinoma is the most frequent pituitary tumor among women of childbearing age. Fewer studies have addressed the outcome of prolactinomas after gestation. Objective: The aim was to study the spontaneous remission rate and change in tumor size after pregnancy and/or lactation in women with prolactinomas. Patients and Methods: Retrospective study conducted at a tertiary care center of north India. Records of 25 women with 31 pregnancies (20 microprolactinomas and 11 macroprolactinomas), who conceived on dopamine agonist (cabergoline) were studied. Cabergoline was stopped at conception in 24 pregnancies and continued in 7. Serum prolactin was noted 3 months after delivery and/or lactation. Magnetic resonance imaging available at last visit after delivery and/or lactation was also noted. Remission was defined as normal serum prolactin after pregnancy and/or lactation without use of cabergoline. Results: Among patients in whom cabergoline was stopped during pregnancy (n = 24), 41.6% (n = 10) had prolactin in normal range (achieved remission) after pregnancy and/or lactation. In 25% (n = 6) of women, adenoma size decreased by more than 50%, in 33%(n = 8), there was no change in adenoma size, and in 42% (n = 10), decrease in adenoma size was less than 50% after pregnancy and/or lactation. The median duration of cabergoline treatment before pregnancy among patients who achieved remission was 60 months against 24 months in those who did not achieve remission. The median pre-pregnancy adenoma size was 5.5 mm in women with remission against 8 mm in women who did not achieve remission. Conclusion: Pregnancy-induced remission of hyperprolactinemia was seen in 41.6% prolactinomas. Longer duration of dopamine agonist treatment before pregnancy, small pre-pregnancy adenoma size, and lower baseline prolactin were associated with high likelihood of remission, though not statistically significant.