A 3D micro-CT assessment of composition and structure of bone tissue after vertical and horizontal alveolar ridge augmentation using CAD/CAM-customized titanium mesh.

OBJECTIVES: To date, no studies have exploited micro-CT in humans to evaluate bone morphology and structure after bone augmentation with CAD/CAM-customized titanium mesh, in mandible and maxilla. The aim of this study was to assess the composition and microstructure of bone biopsy through micro-CT analysis. MATERIALS AND METHODS: Bone augmentation at both maxillary and mandible sites was performed on 30 patients randomly treated with customized mesh, either alone (M-) or covered with resorbable membrane (M+), in both cases filled 50:50 with autogenous bone and xenograft. After 6 months, biopsies were taken and micro-CT was performed on consecutive 1-mm-thick VOIs from coronal to apical side, measuring tissue volumes, trabecular thickness, spacing, and number. RESULTS: In both groups, irrespective of membrane use, bone tissue (M-: 29.76% vs. M+: 30.84%) and residual graft material (M-: 14.87% vs. M+: 13.11%) values were similar. Differences were site-related (maxillary vs. mandibular) with higher percentage of bone tissue and trabecular density of low-mineralized bone and overall bone in the mandible. CONCLUSIONS: The composition and structure of bone tissue, as assessed by micro-CT after alveolar ridge augmentation using CAD/CAM-customized titanium meshes, showed similar features regardless of whether a collagen membrane was applied.

Changes in human skin composition due to intrinsic aging: a histologic and morphometric study.

Skin represents the main barrier against the external environment, but also plays a role in human relations, as one of the prime determinants of beauty, resulting in a high consumer demand for skincare-related pharmaceutical products. Given the importance of skin aging in both medical and social spheres, the present research aims to characterize microscopic changes in human skin composition due to intrinsic aging (as opposed to aging influenced by external factors) via histological analysis of a photoprotected body region. Samples from 25 autopsies were taken from the periumbilical area and classified into four age groups: group 1 (0-12 years), group 2 (13-25 years), group 3 (26-54 years), and group 4 (≥ 55 years). Different traditional histological (hematoxylin-eosin, Masson's trichrome, orcein, toluidine, Alcian blue, and Feulgen reaction) and immunohistochemical (CK20, CD1a, Ki67, and CD31) stains were performed. A total of 1879 images photographed with a Leica DM3000 optical microscope were morphometrically analyzed using Image ProPlus 7.0 for further statistical analysis with GraphPad 9.0. Our results showed a reduction in epidermis thickness, interdigitation and mitotic indexes, while melanocyte count was raised. Papillary but not reticular dermis showed increased thickness with aging. Specifically, in the papillary layer mast cells and glycosaminoglycans were expanded, whereas the reticular dermis displayed a diminution in glycosaminoglycans and elastic fibers. Moreover, total cellularity and vascularization of both dermises were diminished with aging. This morphometric analysis of photoprotected areas reveals that intrinsic aging significantly influences human skin composition. This study paves the way for further research into the molecular basis underpinning these alterations, and into potential antiaging strategies.

Histopathological impact of SARS-CoV-2 on the liver: Cellular damage and long-term complications.

Coronavirus disease 2019 (COVID-19), caused by the highly pathogenic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), primarily impacts the respiratory tract and can lead to severe outcomes such as acute respiratory distress syndrome, multiple organ failure, and death. Despite extensive studies on the pathogenicity of SARS-CoV-2, its impact on the hepatobiliary system remains unclear. While liver injury is commonly indicated by reduced albumin and elevated bilirubin and transaminase levels, the exact source of this damage is not fully understood. Proposed mechanisms for injury include direct cytotoxicity, collateral damage from inflammation, drug-induced liver injury, and ischemia/hypoxia. However, evidence often relies on blood tests with liver enzyme abnormalities. In this comprehensive review, we focused solely on the different histopathological manifestations of liver injury in COVID-19 patients, drawing from liver biopsies, complete autopsies, and in vitro liver analyses. We present evidence of the direct impact of SARS-CoV-2 on the liver, substantiated by in vitro observations of viral entry mechanisms and the actual presence of viral particles in liver samples resulting in a variety of cellular changes, including mitochondrial swelling, endoplasmic reticulum dilatation, and hepatocyte apoptosis. Additionally, we describe the diverse liver pathology observed during COVID-19 infection, encompassing necrosis, steatosis, cholestasis, and lobular inflammation. We also discuss the emergence of long-term complications, notably COVID-19-related secondary sclerosing cholangitis. Recognizing the histopathological liver changes occurring during COVID-19 infection is pivotal for improving patient recovery and guiding decision-making.

Comparative analysis of genetic variants in pleural fluids and solid tissue biopsies of pleural mesothelioma patients: Implications for molecular heterogeneity assessment.

Objectives: This study aims to determine whether the sequencing of DNA extracted from pleural fluids (PFs) of Pleural Mesothelioma (PM) patients accurately represents the genetic information obtained from the solid tissue counterpart biopsies with particular attention to the identification of single nucleotide variants (SNVs). Materials and methods: Single pleural biopsy, PFs, and blood were collected from PM patients. DNA was extracted from these samples and then subjected to Whole-Exome Sequencing. Results: A higher number of SNVs was identified in PFs than in solid tissue biopsies (STBs). Most SNVs were detected in PFs samples but not in STBs samples, while only a few SNVs were detected in STBs samples but not in PFs samples. Conclusion: The current findings support the notion that PFs might offer a more robust depiction of cancer's molecular diversity. Nonetheless, the current outcomes challenge the assertion that liquid biopsies can encompass the entirety of intra-patient variations. Indeed, a subset of potential cancer-driver SNVs was exclusively identified in STBs. However, relying solely on STBs would have precluded the detection of significant SNVs that were exclusively present in PFs. This implies that while PFs serve as a valuable complement to STBs, they do not supplant them.

Exosomes as novel tools for renal cell carcinoma therapy, diagnosis, and prognosis.

Background: Renal Cell Carcinoma (RCC) stands as a formidable challenge within the field of oncology, despite considerable research endeavors. The advanced stages of this malignancy present formidable barriers to effective treatment and management. Objective: This review aims to explore the potential of exosomes in addressing the diagnostic and therapeutic challenges associated with RCC. Specifically, it investigates the role of exosomes as biomarkers and therapeutic vehicles in the context of RCC management. Methods: For this review article, a comprehensive literature search was conducted using databases such as PubMed, employing relevant keywords to identify research articles pertinent to the objectives of the review. Initially, 200 articles were identified, which underwent screening to remove duplicates and assess relevance based on titles and abstracts, followed by a detailed examination of full texts. From the selected articles, relevant data were extracted and synthesized to address the review's objectives. The conclusions were drawn based on a thorough analysis of the findings. The quality was ensured through independent review and resolution of discrepancies among multiple reviewers. Results: Exosomes demonstrate potential as diagnostic tools for early detection, prognosis, and treatment monitoring in RCC. Their ability to deliver various therapeutic agents, such as small interfering RNAs, lncRNAs, chemotherapeutic drugs, and immune-stimulating agents, allows for a personalized approach to RCC management. By leveraging exosome-based technologies, precision and efficacy in treatment strategies can be significantly enhanced. Conclusion: Despite the promising advancements enabled by exosomes in the management of RCC, further research is necessary to refine exosome-based technologies and validate their efficacy, safety, and long-term benefits through rigorous clinical trials. Embracing exosomes as integral components of RCC diagnosis and treatment represents a significant step towards improving patient outcomes and addressing the persistent challenges posed by this malignancy in the field of oncology.

Intracerebellar administration of the chemokine Cxcl3 reduces the volume of medulloblastoma lesions at an advanced stage by promoting the migration and differentiation of preneoplastic precursor cells.

The prognosis for many pediatric brain tumors, including cerebellar medulloblastoma (MB), remains dismal but there is promise in new therapies. We have previously generated a mouse model developing spontaneous MB at high frequency, Ptch1+/-/Tis21-/-. In this model, reproducing human tumorigenesis, we identified the decline of the Cxcl3 chemokine in cerebellar granule cell precursors (GCPs) as responsible for a migration defect, which causes GCPs to stay longer in the proliferative area rather than differentiate and migrate internally, making them targets of transforming insults. We demonstrated that 4-week Cxcl3 infusion in cerebella of 1-month-old mice, at the initial stage of MB formation, forces preneoplastic GCPs (pGCPs) to leave lesions and differentiate, with a complete suppression of MB development. In this study, we sought to verify the effect of 4-week Cxcl3 treatment in 3-month-old Ptch1+/-/Tis21-/- mice, when MB lesions are at an advanced, irreversible stage. We found that Cxcl3 treatment reduces tumor volumes by sevenfold and stimulates the migration and differentiation of pGCPs from the lesion to the internal cerebellar layers. We also tested whether the pro-migratory action of Cxcl3 favors metastases formation, by xenografting DAOY human MB cells in the cerebellum of immunosuppressed mice. We showed that DAOY cells express the Cxcl3 receptor, Cxcr2, and that Cxcl3 triggers their migration. However, Cxcl3 did not significantly affect the frequency of metastases or the growth of DAOY-generated MBs. Finally, we mapped the expression of the Cxcr2 receptor in human MBs, by evaluating a well-characterized series of 52 human MBs belonging to different MB molecular subgroups. We found that Cxcr2 was variably expressed in all MB subgroups, suggesting that Cxcl3 could be used for therapy of different MBs.

Unlocking Precise Lung Cancer Detection Through Minimal Panel Immunostaining in Small Biopsy Samples.

Introduction Lung cancer diagnosis faces challenges due to morphological heterogeneity and limited biopsy tissue. This study evaluates the efficacy of a minimal panel immunostaining technique using immunohistochemical markers like napsin A, thyroid transcription factor 1 (TTF-1), p63, and synaptophysin to improve the precision of lung carcinoma subclassification. Methods A retrospective analytical study was conducted at the Histopathology Laboratory of Saveetha Medical College and Hospital, Chennai, from January 2018 to February 2024. A total of 64 lung carcinoma cases were analyzed. Inclusion criteria included biopsy samples from lung lesions with a confirmed diagnosis of lung carcinoma based on histomorphological examination, covering all age groups and both genders. Non-carcinomatous lung lesions were excluded. Clinical data were obtained from the Medical Information Archiving Software (MIAS) database and histopathological examination request forms. Under a light microscope, tissue samples were examined after being fixed in formalin, processed, and stained with hematoxylin and eosin (H&E). Additionally, a minimal panel of immunohistochemical markers, including napsin A, TTF-1, p63, and synaptophysin, was used to subclassify lung carcinomas. Results The age group older than 50 years was the most affected, with a higher incidence in males. Histologically, 49% of cases were adenocarcinoma, 42% were squamous cell carcinoma, and 9% were small cell carcinoma. Immunohistochemistry (IHC) results adjusted these proportions to 54.6% adenocarcinoma, 31.2% squamous cell carcinoma, and 14% small cell carcinoma, showing a 5.6% increase in adenocarcinoma cases. The most common adenocarcinoma pattern was mixed, followed by acinar. TTF-1 and napsin A were crucial for identifying adenocarcinoma, while p63 was key for squamous cell carcinoma. Synaptophysin confirmed neuroendocrine differentiation in small cell carcinoma. Conclusion Incorporating a minimal panel of IHC markers significantly enhances the accuracy of lung carcinoma subclassification, addressing diagnostic challenges posed by morphological heterogeneity and limited sample size. This approach supports more precise and efficient clinical care for patients with lung cancer. Further validation in diverse clinical settings is recommended.

Pan-cancer evaluation of tumor-infiltrating lymphocytes and programmed cell death protein ligand-1 in metastatic biopsies and matched primary tumors.

Tumor immunological characterization includes evaluation of tumor-infiltrating lymphocytes (TILs) and programmed cell death protein ligand-1 (PD-L1) expression. This study investigated TIL distribution, its prognostic value, and PD-L1 expression in metastatic and matched primary tumors (PTs). Specimens from 550 pan-cancer patients of the SHIVA01 trial (NCT01771458) with available metastatic biopsy and 111 matched PTs were evaluated for TILs and PD-L1. Combined positive score (CPS), tumor proportion score (TPS), and immune cell (IC) score were determined. TILs and PD-L1 were assessed according to PT organ of origin, histological subtype, and metastatic biopsy site. We found that TIL distribution in metastases did not vary according to PT organ of origin, histological subtype, or metastatic biopsy site, with a median of 10% (range: 0-70). TILs were decreased in metastases compared to PT (20% [5-60] versus 10% [0-40], p < 0.0001). CPS varied according to histological subtype (p = 0.02) and biopsy site (p < 0.02). TPS varied according to PT organ of origin (p = 0.003), histological subtype (p = 0.0004), and metastatic biopsy site (p = 0.00004). TPS was higher in metastases than in PT (p < 0.0001). TILs in metastases did not correlate with overall survival. In conclusion, metastases harbored fewer TILs than matched PT, regardless of PT organ of origin, histological subtype, and metastatic biopsy site. PD-L1 expression increased with disease progression. © 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Liquid Biopsies in Follicular Thyroid Carcinomas-A Brief Report.

Thyroid cancer (TC) represents a significant health burden globally, with follicular thyroid cancer (FTC) posing diagnostic challenges despite advancements. This pilot study aimed to evaluate the utility of a liquid biopsy with cell-free DNA (cfDNA) in patients with FTC. Blood samples were collected from 13 patients diagnosed with FTC, DNA extraction was performed, and cfDNA was analyzed using the Illumina's TruSight Oncology 500 High-Throughput panel. The results revealed low tumor mutational burden and minimal pathogenic variants in cfDNA, indicating challenges such as low DNA yield and poor material quality despite adequate coverage. Our findings indicate that cfDNA as an add-on diagnostic tool in patients with FTC might not be a useful supplement.

Heterogeneous Pathological Changes in Liver Lobes During Liver Disease: A Perspective Review.

Liver diseases have a global prevalence of 25%, accounting for 4% of all deaths worldwide, and are associated with a 36% increased risk of fatal and nonfatal cardiovascular events. Metabolic dysfunction-associated steatotic liver disease constitutes the liver expression of metabolic syndrome and represents the primary type of liver disease. Microscopical analysis of biopsies, which allows the evaluation of a small portion of tissue with inferences made to the entire organ, is considered the gold standard for determining the presence of liver diseases. However, potential sampling errors in liver biopsies are conceivable because the obtained tissue represents only a tiny fraction of the entire liver mass and may not accurately reflect the true pathological state. Studies have demonstrated the existence of sampling errors in liver biopsies, particularly concerning the severity of inflammation, degree of fibrosis, and the presence of cirrhosis. Also, clinical studies have shown that histopathological abnormalities are better detected in humans when liver samples are collected from both the right and the left lobes. However, a gap exists in clinical investigation to clarify the role of differences between these lobes in improving the diagnostic and prognostic for liver diseases. Building upon the heterogeneous nature of pathological alterations observed in liver lobes, this perspective review provided recommendations to enhance the precision of diagnosis and prognostic accuracy of liver diseases.

Enough is enough: how many rectal suction biopsies do you need to diagnose Hirschsprung's disease?

PURPOSE: Rectal suction biopsy (RSB) is the gold standard for diagnosing Hirschsprung's disease (HD) in infants. Despite being a common procedure, no standard exists on the number of biopsy specimens and their respective level within the rectum. METHODS: We conducted a retrospective review of epidemiological and pathological data of patients who underwent RSB at our institution between January 2011 and May 2022. During RSB we obtain 4 specimens: at 1 cm, 3 cm and 5 cm above the dentate line, besides one specimen at the dentate line. We used a logistic regression model for statistical analysis and included control variables (e.g. underlying disease, weight at first biopsy, gestational age). RESULTS: A total of 92 patients underwent 115 biopsies, with an average of 3.77 specimens per session. Of the specimens taken at 1 cm above the dentate line 73.9% were conclusive, at 3 cm 75.9% and at 5 cm 79.2%. Specimens taken at the dentate line were squamous or transitional epithelia in 31.5% and therefore of no use for HD diagnostics. The specimen at 3 cm shows the highest discriminative power whether the biopsy session was diagnostic (p-value < 1%). CONCLUSIONS: We propose that a total of three specimens, namely one at 1 cm, one at 3 cm and one at 5 cm above the dentate line, is enough to diagnose or exclude HD.

Biphasic co-detection of melanoma aneuploid tumor cells and tumor endothelial cells in guidance of specifying the field cancerized surgical excision margin and administering immunotherapy.

An optimum safety excision margin (EM) delineated by precise demarcation of field cancerization along with reliable biomarkers that enable predicting and timely evaluating patients' response to immunotherapy significantly impact effective management of melanoma. In this study, optimized biphasic "immunofluorescence staining integrated with fluorescence insitu hybridization" (iFISH) was conducted along the diagnosis-metastasis-treatment-cellular MRD axis to longitudinally co-detect a full spectrum of intact CD31- aneuploid tumor cells (TCs), CD31+ aneuploid tumor endothelial cells (TECs), viable and necrotic circulating TCs (CTCs) and circulating TECs (CTECs) expressing PD-L1, Ki67, p16 and Vimentin in unsliced specimens of the resected primary tumor, EM, dissected sentinel lymph nodes (SLNs) and peripheral blood in an early-stage melanoma patient. Numerous PD-L1+ aneuploid TCs and TECs were detected at the conventional safety EM (2 cm), quantitatively indicating the existence of a field cancerized EM for the first time. Contrary to highly heterogeneous PD-L1 expression and degrees of Chr8 aneuploidy in TCs and TECs in the primary lesions as well as CTCs and CTECs in peripheral blood, almost all TCs and TECs in SLNs and EM were homogeneously PD-L1+ haploid cells. Dynamic monitoring and cellular MRD assessment revealed that, in contrast to PD-L1+ CTCs being responsive to the immune checkpoint inhibitor (ICI-anti-PD-1), multiploid (≥pentasomy 8) PD-L1+ and Ki67+ CTECs were respectively resistant to ICI-sensitized T cells. In therapeutically stressed lymphatic and hematogenous metastatic cascades, stratified phenotypic and karyotypic profiling of iFISH tissue and liquid biopsied TCs, TECs, CTCs and CTECs in future large-cohort studies will enable appropriate re-specification of the optimal safety EM and distribution mapping of in-depth characterized, subcategorized target cells to help illustrate their metastatic relevance, ultimately improving risk stratification and clinical intervention of tumor progression, metastases, therapy resistance and cancer relapse.

Molecular Characterization of Antimicrobial Resistance and Virulence Genotyping among Helicobacter pylori-Positive Dyspeptic Patients in North Iran.

BACKGROUND: Iran has a relatively high prevalence of H. pylori, which correlates with high-risk areas for gastric cancer worldwide. METHODS: Our study aimed to investigate the underlying genetic mechanisms associated with resistance to metronidazole (frxA, rdxA), clarithromycin (23S rRNA), tetracycline (16S rRNA), and fluoroquinolone (gyrA) in H. pylori-positive dyspeptic patients using PCR and sequencing. We further examined the potential correlation between resistance profiles and various virulence genotypes. RESULTS: The rates of genetic mutations associated with resistance to metronidazole, fluoroquinolone, clarithromycin, and tetracycline were found to be 68%, 32.1%, 28.4%, and 11.1%, respectively. Well-documented multiple antibiotic resistance mutations were detected, such as rdxA and frxA (with missense and frameshift alterations), gyrA (Asp91, Asn87), 23S rRNA (A2142G, A2143G), and 16S rRNA (triple-base-pair substitutions AGA926-928→TTC). The cagA+ and vacA s1/m1 types were the predominant genotypes in our study. With the exception of metronidazole and tetracycline, no significant correlation was observed between the cagA+ and cagL+ genotypes and resistance-associated mutations. CONCLUSION: The prevalence of antibiotic resistance-associated mutations in H. pylori was remarkably high in this region, particularly to metronidazole, ciprofloxacin, and clarithromycin. By conducting a simultaneous screening of virulence and resistance genotypes, clinicians can make informed decisions regarding the appropriate therapeutic regimen to prevent the escalation of antibiotic resistance against H. pylori infection in this specific geographical location.

Internalisation of immunoglobulin light chains by cardiomyocytes in AL amyloidosis: what can biopsies tell us?

BACKGROUND: Cardiac involvement in systemic light chain amyloidosis (AL) leads to chronic heart failure and is a major prognosis factor. Severe cellular defects are provoked in cardiac cells by tissue-deposited amyloid fibrils of misfolded free immunoglobulin light chains (LCs) and their prefibrillar oligomeric precursors. OBJECTIVE: Understanding the molecular mechanisms behind cardiac cell cytotoxicity is necessary to progress in therapy and to improve patient management. One key question is how extracellularly deposited molecules exert their toxic action inside cardiac cells. Here we searched for direct evidence of amyloid LC uptake by cardiomyocytes in patient biopsies. METHODS: We immunolocalized LCs in cardiac biopsies from four AL cardiac amyloidosis patients and analysed histopathological images by high resolution confocal microscopy and 3D image reconstruction. RESULTS: We show, for the first time directly in patient tissue, the presence of LCs inside cardiomyocytes, and report their proximity to nuclei and to caveolin-3-rich areas. Our observations point to macropinocytosis as a probable mechanism of LC uptake. CONCLUSIONS: Internalisation of LCs occurs in patient cardiomyocytes. This event could have important consequences for the pathogenesis of the cardiac disease by enabling interactions between amyloid molecules and cellular organelles inducing specific signalling pathways, and might bring new insight regarding treatment.

Antibody Mediated Rejection and T-cell Mediated Rejection Molecular Signatures Using Next-Generation Sequencing in Kidney Transplant Biopsies.

Recently, interest in transcriptomic assessment of kidney biopsies has been growing. This study investigates the use of NGS to identify gene expression changes and analyse the pathways involved in rejection. An Illumina bulk RNA sequencing on the polyadenylated RNA of 770 kidney biopsies was conducted. Differentially-expressed genes (DEGs) were determined for AMR and TCMR using DESeq2. Genes were segregated according to their previous descriptions in known panels (microarray or the Banff Human Organ Transplant (B-HOT) panel) to obtain NGS-specific genes. Pathway enrichment analysis was performed using the Reactome and Kyoto Encyclopaedia of Genes and Genomes (KEGG) public repositories. The differential gene expression using NGS analysis identified 6,141 and 8,478 transcripts associated with AMR and TCMR. While most of the genes identified were included in the microarray and the B-HOT panels, NGS analysis identified 603 (9.8%) and 1,186 (14%) new specific genes. Pathways analysis showed that the B-HOT panel was associated with the main immunological processes involved during AMR and TCMR. The microarrays specifically integrated metabolic functions and cell cycle progression processes. Novel NGS-specific based transcripts associated with AMR and TCMR were discovered, which might represent a novel source of targets for drug designing and repurposing.

Evaluation of Turnaround Times of Diagnostic Biopsies: A Metric of Quality in Surgical Pathology.

Introduction. Timely and accurate diagnosis of diseases is crucial for effective patient care. Turnaround time (TAT) in surgical pathology, defined as the time between accessioning the sample and reporting results, is a key performance indicator reflecting quality and efficiency. This study explores factors affecting TAT for diagnostic biopsies in a tertiary oncology hospital. Methods. A 1-month pilot study was conducted, focusing on 695 in-house diagnostic biopsies. Biopsies were categorized as routine (requiring only hematoxylin and eosin (H&E) staining) or complex cases (requiring additional tests). TAT was defined as the time between sample accessioning and report availability in the electronic medical record, with delays defined as exceeding 3 days for routine cases and 4 days for complex cases. Survival analysis using Kaplan-Meier plots was utilized to analyze TAT. Results. The overall mean TAT was 3.7 ± 2 days, with routine cases at 3.1 ± 2 days and complex cases at 4.8 ± 2 days (P < 0.001). Survival analysis revealed prolonged TAT for complex cases. Organ-specific analysis highlighted variations in TAT, with brain biopsies presenting the highest complexity and longest TAT. Surprisingly, malignant cases demonstrated slightly shorter TATs compared to benign cases (P = 0.026). Delays were observed in 34% of all cases. Conclusions. Laboratory TAT is crucial and is frequently used as a performance benchmark. We analyzed the various causes of delayed TAT in our hospital's histopathology department, with an emphasis on variables in the analytical phase. The results of this study demonstrate that cases involving ancillary techniques had significantly longer TATs compared to routine H&E cases.

Cholesterol Embolic Renal Disease: Experience of a Large Healthcare System.

Context. To determine the clinical-pathological features associated with cholesterol embolic renal disease, and review of literature. Design. This retrospective case series includes patients with cholesterol embolic renal disease (10 of 3087 kidney biopsies) who were diagnosed at Northwell Health, New York, between July 2017 and October 2022. Results. Cholesterol embolic renal disease is a rare disease with a prevalence of 0.32%. Four of 10 patients had intravascular interventional radiology procedures within 6 months prior to kidney biopsy. Four patients had remote interventional radiology history (6 months to 4 years). Seven patients presented with acute kidney injury; 3 patients underwent renal biopsy due to proteinuria. The average baseline creatinine was 2.0 ± 0.9 mg/dL; the creatinine at kidney biopsy and at follow-up was 4.3 ± 3.0 mg/dL and 2.8 ± 1.3 mg/dL, respectively. Eight patients had elevated serum eosinophil counts. Three patients died (mortality rate 30%) in an average follow-up of 4 months (range: 1-10 months). One patient progressed to end-stage kidney disease. The presence of cholesterol clefts is a hallmark of atherosclerotic emboli. Cholesterol clefts were present on the specimen for light microscopy (H&E and special stains) in 9 biopsies; 7 patients had cholesterol clefts in vascular lumens and/or walls. Cholesterol clefts were present on semi-thin sections for electron microscopy examination in 4 biopsies. One patient had cholesterol clefts present in semi-thin sections only. Conclusions. The clinical manifestation of cholesterol embolic renal disease in patients without recent intravascular interventional radiology history can be indolent but is associated with high mortality. Careful examination of all levels with light microscopy, including the perirenal tissue, and semi-thin sections can increase the detection rate of cholesterol embolic renal disease.

High Sensitivity and Specificity Platform to Validate MicroRNA Biomarkers in Cancer and Human Diseases.

We developed a technology for detecting and quantifying trace nucleic acids using a bracketing protocol designed to yield a copy number with approximately ± 20% accuracy across all concentrations. The microRNAs (miRNAs) let-7b, miR-15b, miR-21, miR-375 and miR-141 were measured in serum and urine samples from healthy subjects and patients with breast, prostate or pancreatic cancer. Detection and quantification were amplification-free and enabled using osmium-tagged probes and MinION, a nanopore array detection device. Combined serum from healthy men (Sigma-Aldrich, St. Louis, MO, USA #H6914) was used as a reference. Total RNA isolated from biospecimens using commercial kits was used as the miRNA source. The unprecedented ± 20% accuracy led to the conclusion that miRNA copy numbers must be normalized to the same RNA content, which in turn illustrates (i) independence from age, sex and ethnicity, as well as (ii) equivalence between serum and urine. miR-21, miR-375 and miR-141 copies in cancers were 1.8-fold overexpressed, exhibited zero overlap with healthy samples and had a p-value of 1.6 × 10-22, tentatively validating each miRNA as a multi-cancer biomarker. miR-15b was confirmed to be cancer-independent, whereas let-7b appeared to be a cancer biomarker for prostate and breast cancer, but not for pancreatic cancer.

Activation of macroautophagy and chaperone-mediated autophagy in human skeletal muscle by high-intensity exercise in normoxia and hypoxia and after recovery with or without post-exercise ischemia.

Autophagy is essential for the adaptive response to exercise and physiological skeletal muscle functionality. However, the mechanisms leading to the activation of macroautophagy and chaperone-mediated autophagy in human skeletal muscle in response to high-intensity exercise remain elusive. Our findings demonstrate that macroautophagy and chaperone-mediated autophagy are stimulated by high-intensity exercise in normoxia (PIO2: 143 mmHg) and severe acute hypoxia (PIO2: 73 mmHg) in healthy humans. High-intensity exercise induces macroautophagy initiation through AMPKα phosphorylation, which phosphorylates and activates ULK1. ULK1 phosphorylates BECN1 at Ser15, eliciting the dissociation of BECN1-BCL2 crucial for phagophore formation. Besides, high-intensity exercise elevates the LC3B-II:LC3B-I ratio, reduces total SQSTM1/p62 levels, and induces p-Ser349 SQSTM1/p62 phosphorylation, suggesting heightened autophagosome degradation. PHAF1/MYTHO, a novel macroautophagy biomarker, is highly upregulated in response to high-intensity exercise. The latter is accompanied by elevated LAMP2A expression, indicating chaperone-mediated autophagy activation regardless of post-exercise HSPA8/HSC70 downregulation. Despite increased glycolytic metabolism, severe acute hypoxia does not exacerbate the autophagy signaling response. Signaling changes revert within 1 min of recovery with free circulation, while the application of immediate post-exercise ischemia impedes recovery. Our study concludes that macroautophagy and chaperone-mediated autophagy pathways are strongly activated by high-intensity exercise, regardless of PO2, and that oxygenation is necessary to revert these signals to pre-exercise values. PHAF1/MYTHO emerges as a pivotal exercise-responsive autophagy marker positively associated with the LC3B-II:LC3B-I ratio.