Human Kidney Tissue Research Articles

Fibronectin type III domain containing protein 5/irisin alleviated sepsis-induced acute kidney injury by abating ferroptosis through the adenosine 5'-monophosphate-activated protein kinase/nuclear factor erythroid-2-related factor 2 signaling pathway

Objective: Ferroptosis has been described in association with acute kidney injury (AKI)-induced sepsis. Fibronectin type III domain containing protein 5 (FNDC5)/irisin plays a crucial role in renal protection. The objective of this study was to investigate whether FNDC5/irisin is involved in AKI-induced sepsis by modulating ferroptosis, and the molecular mechanisms that may be involved. Material and Methods: A sepsis-induced AKI model was built in vivo and in vitro through lipopolysaccharide (LPS) intervention. FNDC5, adenosine 5'-monophosphate-activated protein kinase (AMPK), phospho-AMPK (p-AMPK), nuclear factor erythroid-2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), glutathione peroxidase 4 (GPX4), and acyl-CoA synthetase long-chain family member 4 (ACSL4) concentrations in cells and mouse kidney tissues were appraised by Western blot. Pro-inflammatory cytokines concentrations in cell supernatants and mouse kidney tissues were appraised by enzyme-linked immunosorbent assay. Fe2+ concentration in cells and mouse kidney tissue was appraised by kit. The apoptosis rate of cells and mouse kidney tissue was measured by flow cytometry. Automatic biochemical analyzer was to test serum creatinine (SCr) and blood urea nitrogen (BUN). The kidney tissue sections from each groups were observed by hematoxylin and eosin staining. Results: LPS abated FNDC5 concentration in human kidney-2 cells and mouse kidney tissue (P < 0.001). Overexpression of FNDC5 can abated proinflammatory cytokines concentrations in cells and mouse kidney tissue (P < 0.01). Meanwhile, overexpression of FNDC5 can boost GPX4 protein concentration, abate ACSL4 protein, and abate Fe2+ concentration in cells and mouse kidney tissues (P < 0.05). In addition, the overexpression of FNDC5 can reduce the rate of apoptosis (P < 0.01). In vivo experiments showed that FNDC5 overexpression reduced serum BUN and SCr concentrations and alleviated pathological damage in the mouse renal tissues (P < 0.05) and exhibited a certain renal protective effect. FNDC5 overexpression can boost p-AMPK/AMPK, Nrf2, and HO-1 protein concentrations (P < 0.01). Conclusion: FNDC5/irisin improves sepsis-induced acute renal injury by abating ferroptosis through the AMPK/Nrf2 signaling pathway.

Exploring the Spatial Distribution of Interstitial Cells in Kidney Tissue.

Interstitial cells are crucial to the development of kidney structure and function, although the mechanism underlying their role in it remains unclear to date. Our previous study identified cell clusters in human fetal kidney tissue, and we further analyzed the interstitial cell cluster within this context. We extracted the barcoded cDNA from tissue samples and prepared spatial transcriptome libraries. Sequencing data were quality-checked, normalized, and clusters were identified using Seurat. Single-cell and spatial data were integrated using multimodal intersection analysis, and cell types were deconvoluted. DEGs in interstitial cells were identified and functionally annotated using DAVID. CellPhoneDB was used to predict ligand-receptor interactions between cell types. The results of the present study revealed that this cluster of interstitial cells appeared to be scattered in the junction between the cortical and medullary regions. The subsequent Kyoto Encyclopedia of Genes and Genome pathway analysis revealed that the differentially expressed genes (DEGs) in this cluster of interstitial cells were involved in the WNT signaling pathway. The Gene Ontology (GO) analysis revealed that these DEGs were involved in multiple pathways associated with kidney development, with six of the genes (NKD2, TCF21, WNT5A, WNT4, MDK, and SFRP1) associated with kidney development exhibiting significant upregulation. Accordingly, it was inferred that these interstitial cells might be involved in regulating epithelial cell differentiation, ureteral bud development, and morphogenesis. The subsequent cell-cell communication analysis revealed that the cellular crosstalk was primarily regulated mainly by ligand-receptor pairs. Additionally, 17 genes reported to be associated with kidney disease were focused on, and these genes were found to be predominantly expressed in a single-cell type. In summary, the present study revealed the characteristics of a previously identified cluster of interstitial cells in the kidney tissue, thereby providing fresh insights into the process of kidney development.

Optimized protocol for the multiomics processing of cryopreserved human kidney tissue.

Biobanking of tissue from clinically obtained kidney biopsies for later analysis with multiomic approaches, such as single-cell technologies, proteomics, metabolomics, and the different types of imaging, is an inevitable step to overcome the need of disease model systems and toward translational medicine. Hence, collection protocols that ensure integration into daily clinical routines by the usage of preservation media that do not require liquid nitrogen but instantly preserve kidney tissue for both clinical and scientific analyses are necessary. Thus, we modified a robust single-nucleus dissociation protocol for kidney tissue stored snap-frozen or in the preservation media RNAlater and CellCover. Using at first porcine kidney tissue as a surrogate for human kidney tissue, we conducted single-nucleus RNA sequencing with the widely recognized Chromium 10X Genomics platform. The resulting datasets from each storage condition were analyzed to identify any potential variations in transcriptomic profiles. Furthermore, we assessed the suitability of the preservation media for additional analysis techniques such as proteomics, metabolomics, and the preservation of tissue architecture for histopathological examination including immunofluorescence staining. In this study, we show that in daily clinical routines, the preservation medium RNAlater facilitates the collection of highly preserved human kidney biopsies and enables further analysis with cutting-edge techniques like single-nucleus RNA sequencing, proteomics, and histopathological evaluation. Only metabolome analysis is currently restricted to snap-frozen tissue. This work will contribute to build tissue biobanks with well-defined cohorts of the respective kidney disease that can be deeply molecularly characterized, opening up new horizons for the identification of unique cells, pathways and biomarkers for the prevention, early identification, and targeted therapy of kidney diseases.NEW & NOTEWORTHY In this study, we addressed challenges in integrating clinically obtained kidney biopsies into everyday clinical routines. Using porcine kidneys, we evaluated preservation media (RNAlater and CellCover) versus snap freezing for multi-omics processing. Our analyses highlighted RNAlater's suitability for single-nucleus RNA sequencing, proteome analysis and histopathological evaluation. Only metabolomics are currently restricted to snap-frozen biopsies. Our research established a cryopreservation protocol that facilitates tissue biobanking for advancing precision medicine in nephrology.

ACE2 and TMPRSS2 in human kidney tissue and urine extracellular vesicles with age, sex, and COVID-19.

SARS-CoV-2 virus infects cells by engaging with ACE2 requiring protease TMPRSS2. ACE2 is highly expressed in kidneys. Predictors for severe disease are high age and male sex. We hypothesized that ACE2 and TMPRSS2 proteins are more abundant (1) in males and with increasing age in kidney and (2) in urine and extracellular vesicles (EVs) from male patients with COVID-19 and (3) SARS-CoV-2 is present in urine and EVs during infection. Kidney cortex samples from patients subjected to cancer nephrectomy (male/female; < 50years/˃75years, n = 24; ˃80years, n = 15) were analyzed for ACE2 and TMPRSS2 protein levels. Urine from patients hospitalized with SARS-CoV-2 infection was analyzed for ACE2 and TMPRSS2. uEVs were used for immunoblotting and SARS-CoV-2 mRNA and antigen detection. Tissue ACE2 and TMPRSS2 protein levels did not change with age. ACE2 was not more abundant in male kidneys in any age group. ACE2 protein was associated with proximal tubule apical membranes in cortex. TMPRSS2 was observed predominantly in the medulla. ACE2 was elevated significantly in uEVs and urine from patients with COVID-19 with no sex difference compared with urine from controls w/wo albuminuria. TMPRSS2 was elevated in uEVs from males compared to female. ACE2 and TMPRSS2 did not co-localize in uEVs/apical membranes. SARS-CoV-2 nucleoprotein and mRNA were not detected in urine. Higher kidney ACE2 protein abundance is unlikely to explain higher susceptibility to SARS-CoV-2 infection in males. Kidney tubular cells appear not highly susceptible to SARS-CoV-2 infection. Loss of ACE2 into urine in COVID could impact susceptibility and angiotensin metabolism.

SMYD2 Promotes Calcium Oxalate-Induced Glycolysis in Renal Tubular Epithelial Cells via PTEN Methylation.

Background/Objectives: Damage to renal tubular cells (RTCs) represents a critical pathological manifestation in calcium oxalate (CaOx) stone disease, but the underlying mechanism remains elusive. Energy metabolism reprogramming is a vital influencer of RTC survival, and SMYD2 is a histone methylation transferase that has been extensively implicated in various metabolic disorders. Hence, this research aimed to identify whether SMYD2 induces the reprogramming of energy metabolism in RTCs exposed to CaOx nephrolithiasis. Methods: Kidney samples were obtained from patients who underwent laparoscopic nephrectomy for non-functioning kidneys caused by nephrolithiasis. The glyoxylate-induced CaOx stone mice model was established and treated with AZ505. The SMYD2-knockout HK-2 cell line was constructed. Histological changes were evaluated by HE, VK, Tunel, Masson stainings. The molecular mechanism was explored through co-immunoprecipitation and western blotting. Results: The results found that SMYD2 upregulation led to energy reprogramming to glycolysis in human kidney tissue samples and in mice with CaOx nephrolithiasis. We also identified the substantial involvement of glycolysis in the induction of apoptosis, inflammation, and epithelial-mesenchymal transition (EMT) in HK-2 cells caused by calcium oxalate monohydrate (COM). In vivo and in vitro results demonstrated that SMYD2 inhibition reduces glycolysis, kidney injury, and fibrosis. Mechanistically, SMYD2 was found to promote metabolic reprogramming of RTCs toward glycolysis by activating the AKT/mTOR pathway via methylated PTEN, which mediates CaOx-induced renal injury and fibrosis. Conclusions: Our findings reveal an epigenetic regulatory role of SMYD2 in metabolic reprogramming in CaOx nephrolithiasis and associated kidney injury, suggesting that targeting SMYD2 and glycolysis may represent a potential therapeutic strategy for CaOx-induced kidney injury and fibrosis.

Unveiling the role of transgelin as a prognostic and therapeutic target in kidney fibrosis via a proteomic approach

Chronic kidney disease (CKD) progression involves tubulointerstitial fibrosis, a process characterized by excessive extracellular matrix accumulation. To identify potential biomarkers for kidney fibrosis, we performed mass spectrometry-based proteomic profiling of human kidney tubular epithelial cells and kidney tissue from a 5/6 nephrectomy rat model. Multidisciplinary analysis across kidney fibrosis models revealed 351 differentially expressed proteins associated with kidney fibrosis, and they were enriched in processes related to the extracellular matrix, kidney aging, and mitochondrial functions. Network analysis of the selected proteins revealed five crucial proteins, of which transgelin emerged as a candidate protein that interacts with known fibrosis-related proteins. Concordantly, the gene expression of transgelin in the kidney tissue from the 5/6 nephrectomy model was elevated. Transgelin expression in kidney tissue gradually increased from intermediate to advanced fibrosis stages in 5/6 Nx rats and mice with unilateral ureteral obstruction. Subsequent validation in kidney tissue and urine samples from patients with CKD confirmed the upregulation of transgelin, particularly under advanced disease stages. Moreover, we investigated whether blocking TAGLN ameliorated kidney fibrosis and reduced reactive oxygen species levels in cellular models. In conclusion, our proteomic approach identified TAGLN as a potential noninvasive biomarker and therapeutic target for CKD-associated kidney fibrosis, suggesting its role in modulating mitochondrial dysfunction and oxidative stress responses.

Spatial-omic Profiling of Human Kidney Tissues Using SeqStain Stratifies Disease Pathology

Spatial-omic Profiling of Human Kidney Tissues Using SeqStain Stratifies Disease Pathology

Human Kidney Tissue Proteomics Unveils Complement Factor D Involved in the Pathogenesis of Focal Segmental Glomerulosclerosis

Human Kidney Tissue Proteomics Unveils Complement Factor D Involved in the Pathogenesis of Focal Segmental Glomerulosclerosis

Optimized Protocol for the Multiomics Processing of Cryopreserved Human Kidney Tissue

Optimized Protocol for the Multiomics Processing of Cryopreserved Human Kidney Tissue

Accumulation of 3-Monochloro-Propanediol Esters in Kidney Tissues of Patients with Human Renal Cell Carcinoma

Background: 3-Monochloro-propanediol esters (3-MCPDEs), commonly found in refined edible oils and related products, have generated concerns due to their nephrotoxicity and carcinogenicity, yet clinical evidence remains limited. Objectives: In this study, we aimed to assess, for the first time, the accumulation of 3-MCPDEs in human kidney tissues, focusing on 68 participants, some with and others without renal cell carcinoma (RCC). Methods: An analytical method for 3-MCPDE determination in kidney tissues underwent partial validation to ensure its suitability for sample analysis. The analyst was blind to the sample groups. Results: Results revealed significantly higher 3-MCPDE levels in RCC patients compared to non-RCC counterparts (0.22 vs. 0.01 µg/g) (p &lt; 0.01). Moreover, no significant correlation was found between 3-MCPDE levels and tumor stage or size in the RCC group. Conclusions: Accumulation of 3-MCPDEs in humans, with significantly higher levels was observed in kidney tumor specimens compared to non-patients. These findings suggest minimizing the intake of 3-MCPD and its esters in diets in order to reduce potential negative health impacts.

Exploring immune response toward transplanted human kidney tissues assembled from organoid building blocks

The increasing scarcity of organs and the significant morbidity linked to dialysis require the development of engineered kidney tissues from human-induced pluripotent stem cells. Integrative approaches that synergize scalable kidney organoid differentiation, tissue biomanufacturing, and comprehensive assessment of their immune response and host integration are essential to accomplish this. Here, we create engineered human kidney tissues composed of organoid building blocks (OBBs) and transplant them into mice reconstituted with allogeneic human immune cells. Tissue-infiltrating human immune cells are composed of effector Tcells and innate cells. This immune infiltration leads to kidney tissue injury characterized by reduced microvasculature, enhanced kidney cell apoptosis, and an inflammatory gene signature comparable to kidney organ transplant rejection in humans. Upon treatment with the immunosuppressive agent rapamycin, the induced immune response is greatly suppressed. Our model is a translational platform to study engineered kidney tissue immunogenicity and develop therapeutic targets for kidney rejection.

CFAP47 is implicated in X-linked polycystic kidney disease

IntroductionAutosomal dominant polycystic kidney disease (ADPKD) is a well-described condition in which ∼80% of all cases have a genetic explanation, and among sporadic cases without a family history, the genetic bases remains unclear in ∼30% of cases. This study aimed to identify genes associated with polycystic kidney disease (PKD) in patients with sporadic cystic kidney disease in which a clear genetic change was not identified in established genes. MethodsA next-generation sequencing panel analyzed known genes related to kidney cysts in 118 sporadic cases, followed by whole-genome sequencing on 47 unrelated individuals without identified candidate variants. Immunohistological examination was then conducted on both human kidney tissue and kidneys from Cfap47-/Y mice. ResultsThree male patients were found to have rare missense variants in the X-linked gene Cilia And Flagella Associated Protein 47 (CFAP47), none of whom had a family history of the condition. CFAP47 was expressed in primary cilia of human kidney tubules, and knockout mice exhibited vacuolation of tubular cells and tubular dilation, providing evidence that CFAP47 is a causative gene involved in cyst formation. ConclusionThis discovery of CFAP47 as a newly identified gene associated with PKD, displaying X-linked inheritance, emphasizes the need for further cases to understand the role of CFAP47 in PKD.

Abstract P367: Cross-Species Epigenomic Maps of Intergenic Loci for Hypertension and Blood Pressure

Many non-coding SNPs identified in Genome-Wide Association Studies (GWAS) likely affect BP-related gene expression through epigenetic mechanisms. This study analyzes the comparative genomic and epigenomic landscapes in human and rat kidney tissues, including kidney proximal tubule (PT) and medullary thick ascending limb (mTAL), focusing on the major challenge of identifying conserved regulatory elements in intergenic regions. The purpose of our study is to identify BP gene regulatory elements in intergenic regions that are conserved from human to rat for in vivo validation and mechanistic studies. Our ultimate goal is to generate high-resolution, genome-wide epigenomic maps of key BP-relevant tissues, using ATAC-seq, Hi-C, CUT&amp;Tag, DNA methylation profiling and RNA-seq. We are creating visualization hubs for human and rat, integrating our data within the UCSC Genome Browser environment. UCSC contains extensive data for humans but very little for rats, particularly for the updated genome assemblies. To address this gap, we integrated our results with data from the Rat Genome Database (RGD). To date, we harmonized ATAC-seq data from human and rat PT and mTAL and displayed these data in RGD’s JBrowse2 genome browser for unique comparative genome views. Based on our initial analysis of data density and intensity, we are developing a novel pattern recognition algorithm to identify epigenomic marks that are conserved across rat and human genomes. Existing algorithms predominantly focus on the conservation of coding genes, gene families, and genomic sequences. Here, average normalized peak intensities for ATAC-seq data across tissues were subject to canonical correlation between human and rat to identify conserved epigenomic patterns within the BP GWAS loci. Here we present an example of a human BP locus in the intergenic region between NPR3 and TARS1 on chromosome 5 which has a region of conserved synteny with rat chromosome 2. Comparative analysis and visualization of this region show significant correlation between ATAC-seq peaks from rat and human PT and mTAL at this locus, with predicted conserved regulatory elements. These visualizations can uncover novel biological insights and identify potential regulatory targets for intervention. This work underscores the importance of advanced visualization techniques in understanding hypertension pathogenesis, facilitating cross-species comparisons, and enhancing the utility of genomic databases.

MiRNA and mRNA Signatures in Human Acute Kidney Injury Tissue

Acute kidney injury (AKI) is an important contributor to the development of chronic kidney disease (CKD). There is a need to understand molecular mediators that drive recovery and progression to CKD. In particular, the regulatory role of miRNAs in AKI is poorly understood. miRNA and mRNA sequencing were performed on biobanked human kidney tissues obtained in the routine care of subjects with a diagnosis of AKI, minimal change disease (MCD), or without known kidney disease in nephrectomy (Ref) tissue. mRNA analysis revealed that Ref tissues exhibited an injury signature similar to AKI, and not identified in MCD samples. The transcriptomic signature of human AKI was enriched in pathways involved in cell adhesion, epithelial-to-mesenchymal transition, and cell cycle arrest (e.g., CDH6, ITGB6, CDKN1A). In AKI, upregulation of miR-146a, miR-155, miR-142, miR-122 was associated with pathways involved in immune cell recruitment, inflammation, and epithelial-to-mesenchymal transition. miR-122 and miR-146 are associated with downregulation of DDR2 and IGFBP6, genes involved in recovery and progression of kidney disease. These data provide integrated miRNA signatures that complement mRNA and other epigenetic data available in kidney atlases.

P02-35 3D primary human kidney tissue model for nephrotoxicity

P02-35 3D primary human kidney tissue model for nephrotoxicity

MiR-148b-5p regulates hypercalciuria and calcium-containing nephrolithiasis

Calcium-containing stones represent the most common form of kidney calculi, frequently linked to idiopathic hypercalciuria, though their precise pathogenesis remains elusive. This research aimed to elucidate the molecular mechanisms involved by employing urinary exosomal microRNAs as proxies for renal tissue analysis. Elevated miR-148b-5p levels were observed in exosomes derived from patients with kidney stones. Systemic administration of miR-148b-5p in rat models resulted in heightened urinary calcium excretion, whereas its inhibition reduced stone formation. RNA immunoprecipitation combined with deep sequencing identified miR-148b-5p as a suppressor of calcitonin receptor (Calcr) expression, thereby promoting urinary calcium excretion and stone formation. Mice deficient in Calcr in distal epithelial cells demonstrated elevated urinary calcium excretion and renal calcification. Mechanistically, miR-148b-5p regulated Calcr through the circRNA-83536/miR-24-3p signaling pathway. Human kidney tissue samples corroborated these results. In summary, miR-148b-5p regulates the formation of calcium-containing kidney stones via the circRNA-83536/miR-24-3p/Calcr axis, presenting a potential target for novel therapeutic interventions to prevent calcium nephrolithiasis.Graphical abstract

Unbound Fractions of PFAS in Human and Rodent Tissues: Rat Liver a Suitable Proxy for Evaluating Emerging PFAS?

Adverse health effects associated with exposures to perfluoroalkyl and polyfluoroalkyl substances (PFAS) are a concern for public health and are driven by their elimination half-lives and accumulation in specific tissues. However, data on PFAS binding in human tissues are limited. Accumulation of PFAS in human tissues has been linked to interactions with specific proteins and lipids in target organs. Additional data on PFAS binding and unbound fractions (funbound) in whole human tissues are urgently needed. Here, we address this gap by using rapid equilibrium dialysis to measure the binding and funbound of 16 PFAS with 3 to 13 perfluorinated carbon atoms (ηpfc = 3-13) and several functional headgroups in human liver, lung, kidney, heart, and brain tissue. We compare results to mouse (C57BL/6 and CD-1) and rat tissues. Results show that funbound decreases with increasing fluorinated carbon chain length and hydrophobicity. Among human tissues, PFAS binding was generally greatest in brain > liver ≈ kidneys ≈ heart > lungs. A correlation analysis among human and rodent tissues identified rat liver as a suitable surrogate for predicting funbound for PFAS in human tissues (R2 ≥ 0.98). The funbound data resulting from this work and the rat liver prediction method offer input parameters and tools for toxicokinetic models for legacy and emerging PFAS.

Spatial proteomics of human diabetic kidney disease, from health to class III.

Diabetic kidney disease (DKD) is the leading cause of chronic and end-stage kidney disease in the USA and worldwide. Animal models have taught us much about DKD mechanisms, but translation of this knowledge into treatments for human disease has been slowed by the lag in our molecular understanding of human DKD. Using our Spatial TissuE Proteomics (STEP) pipeline (comprising curated human kidney tissues, multiplexed immunofluorescence and powerful analysis tools), we imaged and analysed the expression of 21 proteins in 23 tissue sections from individuals with diabetes and healthy kidneys (n=5), compared to those with DKDIIA, IIA-B and IIB (n=2 each) and DKDIII (n=1). These analyses revealed the existence of 11 cellular clusters (kidney compartments/cell types): podocytes, glomerular endothelial cells, proximal tubules, distal nephron, peritubular capillaries, blood vessels (endothelial cells and vascular smooth muscle cells), macrophages, myeloid cells, other CD45+ inflammatory cells, basement membrane and the interstitium. DKD progression was associated with co-localised increases in inflammatory cells and collagen IV deposition, with concomitant loss of native proteins of each nephron segment. Cell-type frequency and neighbourhood analyses highlighted a significant increase in inflammatory cells and their adjacency to tubular and αSMA+ (α-smooth muscle actin-positive) cells in DKD. Finally, DKD progression showed marked regional variability within single tissue sections, as well as inter-individual variability within each DKD class. Using the STEP pipeline, we found alterations in protein expression, cellular phenotypic composition and microenvironment structure with DKD progression, demonstrating the power of this pipeline to reveal the pathophysiology of human DKD.

Multiscale topology classifies cells in subcellular spatial transcriptomics

Spatial transcriptomics measures in situ gene expression at millions of locations within a tissue1, hitherto with some trade-off between transcriptome depth, spatial resolution and sample size2. Although integration of image-based segmentation has enabled impactful work in this context, it is limited by imaging quality and tissue heterogeneity. By contrast, recent array-based technologies offer the ability to measure the entire transcriptome at subcellular resolution across large samples3–6. Presently, there exist no approaches for cell type identification that directly leverage this information to annotate individual cells. Here we propose a multiscale approach to automatically classify cell types at this subcellular level, using both transcriptomic information and spatial context. We showcase this on both targeted and whole-transcriptome spatial platforms, improving cell classification and morphology for human kidney tissue and pinpointing individual sparsely distributed renal mouse immune cells without reliance on image data. By integrating these predictions into a topological pipeline based on multiparameter persistent homology7–9, we identify cell spatial relationships characteristic of a mouse model of lupus nephritis, which we validate experimentally by immunofluorescence. The proposed framework readily generalizes to new platforms, providing a comprehensive pipeline bridging different levels of biological organization from genes through to tissues.

Injection of luteinizing hormone or human chorionic gonadotropin increases calcium excretion and serum PTH in males

Animal and human studies have suggested that sex steroids have calciotropic actions, and it has been proposed that follicle-stimulating hormone (FSH) may exert direct effects on bone. Here, we demonstrate the expression of the receptor for Luteinizing hormone (LH) and human choriogonadotropin (hCG), LHCGR, in human kidney tissue, suggesting a potential influence on calcium homeostasis. To investigate the role of LHCGR agonist on calcium homeostasis in vivo, we conducted studies in male mice and human subjects. Male mice were treated with luteinizing hormone (LH), and human extrapolation was achieved by injecting 5000 IU hCG once to healthy men or men with hypergonadotropic or hypogonadotropic hypogonadism. In mice, LH treatment significantly increased urinary calcium excretion and induced a secondary increase in serum parathyroid hormone (PTH). Similarly, hCG treatment in healthy men led to a significant increase in urinary calcium excretion, serum PTH levels, and 1,25 (OH)2D3, while calcitonin, and albumin levels were reduced, possibly to avoid development of persistent hypocalcemia. Still, the rapid initial decline in ionized calcium coincided with a significant prolongation of the cardiac QTc-interval that normalized over time. The observed effects may be attributed to LH/hCG-receptor (LHCGR) activation, considering the presence of LHCGR expression in human kidney tissue, and the increase in sex steroids occurred several hours after the changes in calcium homeostasis. Our translational study shed light on the intricate relationship between gonadotropins, sex hormones and calcium, suggesting that LHCGR may be influencing calcium homeostasis directly or indirectly.