Herbo-Mineral Medicine, Lithom Exhibits Anti-Nephrolithiasis Activity in Rat Model of Hyperoxaluria by Attenuating Calcium Oxalate Crystal Formation and Oxidative Stress.

BACKGROUND: Calcium oxalate monohydrate (COM) forms the most common type of kidney stones observed in clinics, elevated levels of urinary oxalate being the principal risk factor for such an etiology. The objective of the present study was to evaluate the anti-nephrolithiatic effect of herbo-mineral formulation, Lithom. METHODS: The in vitro biochemical synthesis of COM crystals in the presence of Lithom was performed and observations were made by microscopy and Scanning Electron Microscope (SEM) based analysis for the detection of crystal size and morphology. The phytochemical composition of Lithom was evaluated by Ultra-High-Performance Liquid Chromatography (UHPLC). The in vivo model of Ethylene glycol-induced hyperoxaluria in Sprague-Dawley rats was used for the evaluation of Lithom. The animals were randomly allocated to 5 different groups namely Normal control, Disease control (ethylene glycol (EG), 0.75%, 28 days), Allopurinol (50 mg/kg, q.d.), Lithom (43 mg/kg, b.i.d.), and Lithom (129 mg/kg, b.i.d.). Analysis of crystalluria, oxalate, and citrate levels, oxidative stress parameters (malondialdehyde (MDA), catalase, myeloperoxidase (MPO)), and histopathology by hematoxylin and eosin (H&E) and Von Kossa staining was performed for evaluation of Lithom. RESULTS: The presence of Lithom during COM crystals synthesis significantly reduced the average crystal area, feret's diameter, and area-perimeter ratio, in a dose-dependent manner. SEM analysis revealed that COM crystals synthesized in the presence of 100 and 300 µg/mL of Lithom exhibited a veritable morphological transition from irregular polygons with sharp edges to smoothened smaller cuboid polygons. UHPLC analysis of Lithom revealed the presence of Trigonelline, Bergenin, Xanthosine, Adenosine, Bohoervinone B, Vanillic acid, and Ellagic acid as key phytoconstituents. In EG-induced SD rats, the Lithom-treated group showed a decrease in elevated urinary oxalate levels, oxidative stress, and renal inflammation. Von Kossa staining of kidney tissue also exhibited a marked reduction in crystal depositions in Lithom-treated groups. CONCLUSION: Taken together, Lithom could be a potential clinical-therapeutic alternative for management of nephrolithiasis.

Inhibitory effect of Incarvillea diffusa Royle extract in the formation of calcium oxalate nephrolithiasis by regulating ROS-induced Nrf2/HO-1 pathway in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Calcium oxalate (CaOx) kidney stones are widely acknowledged as the most prevalent type of urinary stones, with high incidence and recurrence rates. Incarvillea diffusa Royle (ID) is a traditionally used medicinal herb in the Miao Minzu of Guizhou province, China, for treating urolithiasis. However, the active components and the underlying mechanism of its pharmacodynamic effects remain unclear. AIM OF THE STUDY: This study aimed to investigate the potential inhibitory effect of the active component of ID on the formation of CaOx nephrolithiasis and elucidate the underlying mechanism. MATERIALS AND METHODS: In vivo, a CaOx kidney stone model was induced in Sprague-Dawley (SD) rats using an ethylene glycol and ammonium chloride protocol for four weeks. Forty-eight male SD rats were randomly assigned to 6 groups (n = 8): blank group, model group, apocynin group, and low, medium, and high dose of ID's active component (IDW) groups. After three weeks of administration, rat urine, serum, and kidney tissues were collected. Renal tissue damage and crystallization, Ox, BUN, Ca2+, CRE, GSH, MDA, SOD contents, and levels of IL-1ß, IL-18, MCP-1, caspase-1, IL-6, and TNF-α in urine, serum, and kidney tissue were assessed using HE staining and relevant assay kits, respectively. Protein expression of Nrf2, HO-1, p38, p65, and Toll-4 in kidney tissues was quantified via Western blot. The antioxidant capacities of major compounds were evaluated through DPPH, O2·-, and ·OH radical scavenging assays, along with their effects on intracellular ROS production in CaOx-induced HK-2 cells. RESULTS: We found that IDW could significantly reduce the levels of CRE, GSH, MDA, Ox, and BUN, and enhancing SOD activity. Moreover, it could inhibit the secretion of TNF-α, IL-1ß, IL-18, MCP-1, caspase-1, and decreased protein expression of Nrf2, HO-1, p38, p65, and Toll-4 in renal tissue. Three major compounds isolated from IDW exhibited promising antioxidant activities and inhibited intracellular ROS production in CaOx-induced HK-2 cells. CONCLUSIONS: IDW facilitated the excretion of supersaturated Ca2+ and decreased the production of Ox, BUN in SD rat urine, and mitigated renal tissue damage by regulating Nrf2/HO-1 signaling pathway. Importantly, the three major compounds identified as active components of IDW contributed to the inhibition of CaOx nephrolithiasis formation. Overall, IDW holds significant potential for treating CaOx nephrolithiasis.

[New Mutation of CYP24A1 in a Case of Idiopathic Infantile Hypercalcemia Diagnosed in Adulthood].

Mutations in the 24-hydroxylase gene CYP24A1 have been recognized as causes of childhood idiopathic hypercalcemia (IIH), a rare disease (incidence <1:1,000,000 live births) characterized by increased vitamin D sensitivity, with symptomatic severe hypercalcemia. IIH was first described in Great Britain two years after the start of a program of vitamin D supplementation in milk for the prevention of rickets, manifesting in about 200 children with severe hypercalcemia, dehydration, growth failure, weight loss, muscle hypotonia, and nephrocalcinosis. The association between the epidemic occurrence of IIH and vitamin D administration was quickly attributed to intrinsic hypersensitivity to vitamin D, and the pathogenic mechanism was recognized in the inactivation of Cytochrome P450 family 24 subfamily A member 1 (CYP24A1), which was identified as the molecular basis of the pathology. The phenotypic spectrum of CYP24A1 mutation can be variable, manifesting predominantly with childhood onset and severe symptomatology (severe hypercalcemia, growth retardation, lethargy, muscle hypotonia, dehydration), but also with juvenile-adult onset forms with nephrolithiasis, nephrocalcinosis, and alterations in phosphocalcium homeostasis. We describe the case of a patient in whom the diagnosis of IIH was made in adulthood, presenting with finding of nephrocalcinosis in childhood, and with subsequent onset of severe hypercalcemia with hypercalciuria, hypoparathyroidism, hypervitaminosis D, and recurrent renal lithiasis. Genetic investigation revealed the presence in homozygosity of the c_428_430delAAG_p.Glu143del variant in the CYP24A1 gene with autosomal recessive transmission, a mutation not reported in the literature.

Nephrolithiasis in two patients on iron chelation therapy: A case report.

Drug-induced nephrolithiasis can arise from insoluble components within medications or crystallization of metabolites due to changes in metabolism and urinary pH. The connection between drugs utilized for iron chelation therapy (ICT) and nephrolithiasis is not well understood. In this report, we describe two pediatric patients diagnosed with nephrolithiasis while undergoing treatment with the chelating agents deferasirox, deferiprone, and deferoxamine for iron overload secondary to repeat blood transfusion.

Network pharmacology and molecular modelling study of Enhydra fluctuans for the prediction of the molecular mechanisms involved in the amelioration of nephrolithiasis.

In view of the ethno medicinal use of Enhydra fluctuans for the treatment of kidney stones; the present study aimed to elucidate the molecular mechanisms involved in the amelioration of nephrolithiasis through a network pharmacology approach. The phytoconstituents were queried in DIGEP-Pred to identify the regulated proteins. The modulated proteins were then enriched in the STRING to predict the protein-protein interactions and the probably regulated pathways were traced in the Kyoto Encyclopedia of Genes and Genomes. Further, the network was constructed using Cytoscape ver 3.5.1. Results showed that ß-carotene was found to be regulating maximum targets i.e. 26. In addition, 63 proteins were triggered by the components in which the vitamin D receptor was targeted by the maximum phytoconstituents i.e. 16. The enrichment analysis identified the regulation of 67 pathways in which fluid shear stress and atherosclerosis-associated pathways (KEGG entry hsa05418) regulated ten genes. Further, protein kinase C-α was traced in 23 different pathways. In addition, the majority of the regulated genes were identified from the extracellular space via the modulation of 43 genes. Also, nuclear receptor activity had the maximum molecular function via the regulation of 7 genes. Likewise, the response to organic substance was predicted to trigger the top genes i.e. 43. In contrast, Stigmasterol, Baicalein-7-o-glucoside, and Kauran-16-ol were found to have a high affinity to bind with the VDR receptor confirmed by the molecular modelling and the dynamics. Hence, the study elucidated the probable molecular mechanisms of E. fluctuans in managing nephrolithiasis and identified the lead molecules, their targets, and possible pathways.Communicated by Ramaswamy H. Sarma.

Novel Calcium-Sensing Receptor (CASR) Mutation in a Family with Autosomal Dominant Hypocalcemia Type 1 (ADH1): Genetic Study over Three Generations and Clinical Characteristics.

INTRODUCTION: Activating mutation of the calcium-sensing receptor gene (CASR) reduces parathyroid hormone secretion and renal tubular reabsorption of calcium, defined as autosomal dominant hypocalcemia type 1 (ADH1). Patients with ADH1 may present with hypocalcemia-induced seizures. Calcitriol and calcium supplementation in symptomatic patients may exacerbate hypercalciuria, leading to nephrocalcinosis, nephrolithiasis, and compromised renal function. METHODS: We report on a family with seven members over three generations with ADH1 due to a novel heterozygous mutation in exon 4 of CASR: c.416T>C. RESULTS: This mutation leads to substitution of isoleucine with threonine in the ligand-binding domain of CASR. HEK293T cells transfected with wild type or mutant cDNAs demonstrated that p.Ile139Thr substitution led to increased sensitivity of the CASR to activation by extracellular calcium relative to the wild-type CASR (EC50 of 0.88 ± 0.02 mM vs. 1.1 ± 0.23 mM, respectively, p < 0.005). Clinical characteristics included seizures (2 patients), nephrocalcinosis and nephrolithiasis (3 patients), and early lens opacity (2 patients). In 3 of the patients, serum calcium and urinary calcium-to-creatinine ratio levels obtained simultaneously over 49 patient-years were highly correlated. Using the age-specific maximal-normal levels of calcium-to-creatinine ratio in the correlation equation, we obtained age-adjusted serum calcium levels that are high enough to reduce hypocalcemia-induced seizures and low enough to reduce hypercalciuria. CONCLUSION: We report on a novel CASR mutation in a three-generation kindred. Comprehensive clinical data enabled us to suggest age-specific upper limit of serum calcium levels, considering the association between serum calcium and renal calcium excretion.

Urinary stone disease in Syrian children.

BACKGROUND: Pediatric urinary stone disease (USD) is a costly medical problem. This study aims to assess the clinical characteristics and outcomes of common and rare causes of pediatric USD. METHODS: A retrospective descriptive cohort study included all children < 13 years of age with confirmed USD admitted to the Children's University Hospital in Damascus, Syria, from January 2013 to December 2019. The study sample was divided into two groups based on etiologies: common and rare causes groups. RESULTS: We evaluated 235 patients; 147 of them were males, and the male-to-female ratio was 1.7:1. The common causes group consisted of 203 patients (mean age 3.52 ± 3.66 years) and mainly included metabolic disorders (45.5%) and anatomical abnormalities (22.3%), while the rare causes group included 32 cases (mean age 4.93 ± 4.08 years), 12 patients with uric acid stones (37.5%), 7 patients with cystinuria (21.9%), and primary hyperoxaluria in 5 patients (15.6%). In addition, 39.6% of study patients were born to consanguineous marriages. Sixty-two patients developed AKI, and eleven patients had chronic kidney disease (CKD). Patients with rare causes were more likely to have AKI, CKD, bilateral stones, and recurrent stones (P-value < 0.05). Stone analysis was performed on 83 patients, and the main stone types were calcium oxalate (34.9%), uric acid (14.4%), and struvite stones (12%). Surgery was the most performed treatment in 101 patients (56.7%). CONCLUSION: Patients with rare causes of pediatric USD are at a higher risk for severe complications and require early diagnosis and management. The high rate of uric acid stones in our society requires further evaluation for possible underlying causes. A higher resolution version of the Graphical abstract is available as Supplementary information.

Gallic acid ameliorates calcium oxalate crystal-induced renal injury via upregulation of Nrf2/HO-1 in the mouse model of stone formation.

BACKGROUND: High prevalence and reoccurrence rate of nephrolithiasis bring about serious socioeconomic and healthcare burden, necessitating the need of effective therapeutic agents. Previous study revealed that gallic acid (GAL) alters the nucleation pathway of calcium oxalate (CaOx). On the other hand, it appears protective role against oxidative injury. Whether GAL could protect against crystal-induced lesion in vivo, and its underlying mechanism is yet unsolved. PURPOSE: This study aims to investigate the protective effects of GAL on the crystal-induced renal injury and its underlying mechanism in the mouse model of stone formation induced by glyoxylic acid. STUDY DESIGN AND METHODS: The mouse model of stone formation was established via successive intraperitoneal injection of glyoxylate. Proximal tubular epithelial cell line HK-2 treated with calcium oxalate monohydrate (COM) was used as in vitro model. The protective role of GAL on nephrolithiasis was tested by determination of tubular injury, crystal deposition and adhesion, levels of inflammatory cytokines, macrophage infiltration and the redox status of kidney. In vitro, effect of GAL on the ROS level and oxidative tubular injury induced by COM were detected, as well as major antioxidant pathway Nrf2/HO-1. RESULTS: Administration of GAL alleviates the renal deposition and adhesion of CaOx stone. Meanwhile, GAL ameliorates the inflammation and renal tubular injury. Level of intracellular ROS, osteopontin and CD44 are reduced, either in the mouse model of stone formation or in the COM-treated HK-2 cells after treatment of GAL. Mechanistically, GAL activates Nrf2/HO-1 pathway in HK-2 cells. Silencing Nrf2 abrogates the protective effect of GAL on the oxidative injury and adhesion of COM in HK-2 cells. CONCLUSION: Taken together, our study demonstrates the protective effect of GAL on the deposition of kidney stone and consequent tubular injury. Induction of the antioxidant pathway Nrf2/HO-1 was found to decrease the level of ROS and oxidative injury, thus implying that GAL could be a potential therapeutic agent for the treatment of nephrolithiasis.

Calcium Citrate Versus Calcium Carbonate in the Management of Chronic Hypoparathyroidism: A Randomized, Double-Blind, Crossover Clinical Trial.

In hypoparathyroidism (HypoPT), calcium supplementation is virtually always required, although the disease is likely to be associated with an increased risk of nephrolithiasis. The use of calcium citrate (Ca-Cit) theoretically could have a positive impact on the nephrolithiasis risk because citrate salts are used to reduce this risk. Our objective was to evaluate the potential therapeutic advantage of Ca-Cit in comparison with calcium carbonate (CaCO3 ) in HypoPT, on nephrolithiasis risk factors, as well as to their ability to maintain desirable serum calcium levels. We also evaluated these preparations on quality of life (QOL). This randomized, double-blind, crossover trial recruited 24 adults with postsurgical chronic hypoparathyroidism at Campus Bio-Medico University of Rome. Participants were randomized 1:1 to Ca-Cit or CaCO3 for 1 month and then crossed over to the other treatment for another month. The primary outcomes were changes in albumin-adjusted serum calcium and in ion activity product of calcium oxalate levels (AP[CaOx] index). Secondary efficacy outcomes included changes in SF-36 survey score, fatigue score, constipation, and adverse events. No difference in terms of AP(CaOx) index was observed between the two groups. However, Ca-Cit was associated with a significant reduction in the oxalate/creatinine ratio compared with CaCO3 (-2.46 mmol/mol [SD 11.93] versus 7.42 mmol/mol [SD 17.63], p = 0.029). Serum calcium and phosphorus concentration was not different between the two calcium preparations. Ca-Cit was associated with less constipation (p = 0.047). No difference was found in QOL scores. Although Ca-Cit did not modify the AP(CaOx) index when compared with CaCO3, it was associated with a reduction in urinary oxalate excretion that could have a potential beneficial effect on nephrolithiasis risk. These results are likely to have clinical implications in HypoPT, particularly those who do not tolerate CaCO3 and those affected by nephrolithiasis. A longer-term experience is needed to confirm these findings. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

The Russian Registry of Chronic Hypoparathyroidism.

INTRODUCTION: Chronic hypoparathyroidism is a relatively rare disease associated with multicomponent medical therapy and various complications. The analysis of large databases of patients with chronic hypoparathyroidism is a necessary tool to enhance quality of medical care, as well as to determine the optimal clinical and therapeutic approaches, and prognostic markers of the disease. THE AIM: of this study is to estimate the clinical and biochemical profile, long-term complications, medical therapy and disease control of the patients with chronic postsurgical and non-surgical hypoparathyroidism. MATERIALS AND METHODS: the cross-sectional, observational, continuous study was based on the Russian Registry of patients with hypoparathyroidism. 544 patients from 63 regions of the Russian Federation were included in this study. RESULTS: The majority of cases had postsurgical etiology (88.4%). Postsurgical hypoparathyroidism prevailed in females (р<0.001). About a half of patients had blood calcium and phosphorus targets, 56 and 52% respectively. Nephrolithiasis was confirmed in 32.5%, nephrocalcinosis - in 12.3% of cases. The risk of nephrocalcinosis/nephrolithiasis increased by 1.85 times with disease duration more than 4.5 years. The cataract was found in 9.4%. The cut-off point for the development of cataracts was 9.5 years, with a 6.96-fold increased risk. The longer duration of hypoparathyroidism of any etiology was associated with more frequent cataract (p=0.0018).We found brain calcification in 4%, arrhythmias in 7.2% and neuropsychiatric symptoms in 5.15% of cases. Generally, the BMD in the studied group corresponded to age values, and there was no evidence for the phenomenon of high bone density. TBS was consistent with normal bone microarchitectonics. In our study, the majority of patients (83.5%) was treated with standard therapy of calcium and vitamin D supplements. 5 patients with severe disease course were treated with rhPTH (1-34). CONCLUSIONS: Analysis of the presented database indicates insufficient diagnosis of the complications associated with chronic hypoparathyroidism. Overall, hypoparathyroidism is associated with higher risks of renal stone formation, decreased GFR, cataract especially in patients with longer duration of disease.

HydroZitLa inhibits calcium oxalate stone formation in nephrolithic rats and promotes longevity in nematode Caenorhabditis elegans.

Low fluid intake, low urinary citrate excretion, and high oxidative stress are main causative factors of calcium oxalate (CaOx) nephrolithiasis. HydroZitLa contains citrate and natural antioxidants and is developed to correct these three factors simultaneously. Antioxidants theoretically can prolong the lifespan of organisms. In this study, we preclinically investigated the antilithogenic, lifespan-extending and anti-aging effects of HydroZitLa in HK-2 cells, male Wistar rats, and Caenorhabditis elegans. HydroZitLa significantly inhibited CaOx crystal aggregation in vitro and reduced oxidative stress in HK-2 cells challenged with lithogenic factors. For experimental nephrolithiasis, rats were divided into four groups: ethylene glycol (EG), EG + HydroZitLa, EG + Uralyt-U, and untreated control. CaOx deposits in kidneys of EG + HydroZitLa and EG + Uralyt-U rats were significantly lower than those of EG rats. Intrarenal expression of 4-hydroxynonenal in EG + HydroZitLa rats was significantly lower than that of EG rats. The urinary oxalate levels of EG + HydroZitLa and EG + Uralyt-U rats were significantly lower than those of EG rats. The urinary citrate levels of EG + HydroZitLa and EG + Uralyt-U rats were restored to the level in normal control rats. In C. elegans, HydroZitLa supplementation significantly extended the median lifespan of nematodes up to 34% without altering feeding ability. Lipofuscin accumulation in HydroZitLa-supplemented nematodes was significantly lower than that of non-supplemented control. Additionally, HydroZitLa inhibited telomere shortening, p16 upregulation, and premature senescence in HK-2 cells exposed to lithogenic stressors. Conclusions, HydroZitLa inhibited oxidative stress and CaOx formation both in vitro and in vivo. HydroZitLa extended the lifespan and delayed the onset of aging in C. elegans and human kidney cells. This preclinical evidence suggests that HydroZitLa is beneficial for inhibiting CaOx stone formation, promoting longevity, and slowing down aging.

Safety and tolerability of high-dose daily vitamin D3 supplementation in the vitamin D and type 2 diabetes (D2d) study-a randomized trial in persons with prediabetes.

BACKGROUND/OBJECTIVES: Routine use of vitamin D supplements has increased substantially in the United States. However, the safety and tolerability of long-term use of high-dose vitamin D are not known. We assessed the safety and tolerability of high-dose, daily vitamin D3 in the vitamin D and type 2 diabetes (D2d) study. SUBJECTS/METHODS: In total, 2423 overweight/obese persons with prediabetes were randomized in a double-blind manner to either 4000 IU of vitamin D3 (the tolerable upper intake level for adults by the National Academy of Medicine) taken daily or matching placebo. All participants were included in this analysis. Incident adverse events (AE) were ascertained 4 times a year at in-person visits (twice a year) and interim remote encounters (twice a year) and were defined as untoward or unfavorable medical occurrences. Serious adverse events (SAE) included death, life-threatening events, and hospitalizations. RESULTS: A total of 8304 AEs occurred during 3 years of follow-up and were less frequent in the vitamin D group compared to placebo (Incidence Rate Ratio [IRR] = 0.94; 95% Confidence Interval (CI) 0.90, 0.98). The overall frequency of protocol-specified AEs of interest, which included nephrolithiasis, hypercalcemia, hypercalciuria, or low estimated glomerular filtration rate, was low and did not differ by group. There were no significant between-group differences in total SAEs (IRR = 0.96 (0.81, 1.14)). CONCLUSION: Vitamin D3 supplementation at 4000 IU per day was safe and well tolerated among overweight/obese participants at high risk for diabetes who were appropriately monitored for safety. In this population, this dose of vitamin D3 did not increase risk of AEs or SAEs, including those previously associated with vitamin D such as hypercalcemia, hypercalciuria, or nephrolithiasis. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov NCT01942694, prospectively registered September 16, 2013.

Dual modulatory effects of diosmin on calcium oxalate kidney stone formation processes: Crystallization, growth, aggregation, crystal-cell adhesion, internalization into renal tubular cells, and invasion through extracellular matrix.

Diosmin is a natural flavone glycoside (bioflavonoid) found in fruits and plants with several pharmacological activities. It has been widely used as a dietary supplement or therapeutic agent in various diseases/disorders. Although recommended, evidence of its protective mechanisms against kidney stone disease (nephrolithiasis/urolithiasis), especially calcium oxalate (CaOx) monohydrate (COM) that is the most common type, remained unclear. In this study, we thus systematically evaluated the effects of diosmin (at 2.5-160 nM) on various stages of kidney stone formation processes, including COM crystallization, crystal growth, aggregation, crystal-cell adhesion, internalization into renal tubular cells and invasion through extracellular matrix (ECM). The results showed that diosmin had dose-dependent modulatory effects on all the mentioned COM kidney stone processes. Diosmin significantly increased COM crystal number and mass during crystallization, but reduced crystal size and growth. While diosmin promoted crystal aggregation, it inhibited crystal-cell adhesion and internalization into renal tubular cells. Finally, diosmin promoted crystal invasion through the ECM. Our data provide evidence demonstrating both inhibiting and promoting effects of diosmin on COM kidney stone formation processes. Based on these dual modulatory activities of diosmin, its anti-urolithiasis role is doubtful and cautions should be made for its use in kidney stone disease.

Theaflavin protects against oxalate calcium-induced kidney oxidative stress injury via upregulation of SIRT1.

Renal tubular cell injury induced by calcium oxalate (CaOx) is a critical initial stage of kidney stone formation. Theaflavin (TF) has been known for its strong antioxidative capacity; however, the effect and molecular mechanism of TF against oxidative stress and injury caused by CaOx crystal exposure in kidneys remains unknown. To explore the potential function of TF on renal crystal deposition and its underlying mechanisms, experiments were conducted using a CaOx nephrocalcinosis mouse model established by glyoxylate intraperitoneal injection, and HK-2 cells were subjected to calcium oxalate monohydrate (COM) crystals, with or without the treatment of TF. We discovered that TF treatment remarkably protected against CaOx-induced kidney oxidative stress injury and reduced crystal deposition. Additionally, miR-128-3p expression was decreased and negatively correlated with SIRT1 level in mouse CaOx nephrocalcinosis model following TF treatment. Moreover, TF suppressed miR-128-3p expression and further abolished its inhibition on SIRT1 to attenuate oxidative stress in vitro. Mechanistically, TF interacted with miR-128-3p and suppressed its expression. In addition, miR-128-3p inhibited SIRT1 expression by directly binding its 3'-untranslated region (UTR). Furthermore, miR-128-3p activation partially reversed the acceerative effect of TF on SIRT1 expression. Taken together, TF exhibits a strong nephroprotective ability to suppress CaOx-induced kidney damage through the recovery of the antioxidant defense system regulated by miR-128-3p/SIRT1 axis. These findings provide novel insights for the prevention and treatment of renal calculus.

Kaempferol alleviates calcium oxalate crystal-induced renal injury and crystal deposition via regulation of the AR/NOX2 signaling pathway.

BACKGROUND: Calcium oxalate (CaOx) crystal deposition and crystal-induced renal tubular epithelial cell injury have been found to fundamentally contribute to the formation of CaOx nephrolithiasis. PURPOSE: In the current work, we aim to study the role and mechanism of kaempferol in CaOx crystal kidney deposition and crystal-induced renal injury. STUDY DESIGN: Mice models and HK-2 cells were used to investigate the effect of kaempferol in CaOx crystal-induced renal injury and crystal deposition in the kidney and its underlying mechanism by a series of experiments. METHODS: CaOx crystal deposition in mice renal tubulars and tubular damage were evaluated. And crystal adhesion to HK-2 cells, as well as cellular injury were identified. Furthermore, the effect of kaempferol on the expression of androgen receptor (AR) in renal tubular epithelial cells was assessed. The interaction between AR and nicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2), and the intrinsic molecular mechanism of how AR regulated NOX2 in HK-2 cells were dissected. Additionally, several different assays were applied to analyze the expression levels of various related genes in this study. RESULTS: It was revealed that kaempferol reduced CaOx crystal deposition in renal tubulars and crystal adhesion to HK-2 cells. Meanwhile, the results of in vivo and in vitro experiments corroborated that crystal-associated cellular injury, oxidative stress, inflammation and over-expression of OPN and CD44 in the kidney were ameliorated by kaempferol. Moreover, kaempferol functioned on inhibiting the expression of AR in renal tubular epithelial cells, and AR was able to up-regulate the expression of NOX2 at the transcriptional level by directly binding to the promoter of NOX2. Kaempferol decreased crystal deposition and crystal-induced renal oxidative and inflammatory injury by the down-regulation of AR/NOX2 signaling pathway. CONCLUSION: Taken together, our study findings suggest that kaempferol has a suppressive effect on renal AR expression, which can attenuate CaOx crystal deposition and crystal-induced kidney injury through repressing oxidative stress and inflammation in the kidney by modulating the AR/NOX2 signaling pathway. It demonstrates that kaempferol may have preventive and therapeutic potential for CaOx nephrolithiasis.

Vitexin exerts protective effects against calcium oxalate crystal-induced kidney pyroptosis in vivo and in vitro.

BACKGROUND: Nephrolithiasis is a common urinary disease with a high recurrence rate of secondary stone formation. Several mechanisms are involved in the onset and recurrence of nephrolithiasis, e.g., oxidative stress, inflammation, apoptosis, and epithelial-mesenchymal transition (EMT). Vitexin, a flavonoid monomer derived from medicinal plants that exert many biological effects including anti-inflammatory and anticancer effects, has not been investigated in nephrolithiasis studies. Moreover, pyroptosis, a form of programmed cell death resulting from inflammasome-associated caspase activation, has not been studied in mice with nephrolithiasis. PURPOSE: We aimed to investigate the protective effect and underlying mechanisms of vitexin in nephrolithiasis, and the related role of pyroptosis in vivo and in vitro. METHODS: Mouse models of nephrolithiasis were established via intraperitoneal injection of glyoxylate, and cell models of tubular epithelial cells and macrophages were established using calcium oxalate monohydrate (COM). Crystal deposition and kidney tissue injury were evaluated by hematoxylin and eosin, and von Kossa staining. Renal oxidative stress indexes including malondialdehyde (MDA), superoxide dismutase (SOD), glutathione (GSH), and catalase (CAT), were analyzed. The renal expression of interleukin-1 beta (IL-1ß), gasdermin D (GSDMD), osteopontin (OPN), CD44, and monocyte chemotactic protein 1 (MCP-1), and EMT-related proteins in renal tubular epithelial cells was assessed. Cell viability and the apoptosis ratio were evaluated. RESULTS: In vivo, vitexin alleviated crystal deposition and kidney tissue injury, and decreased the level of MDA, and increased the levels of SOD, GSH, and CAT. Vitexin also reduced the levels of the pyroptosis-related proteins GSDMD, NLRP3, cleaved caspase-1, and mature IL-1ß, which were elevated in mice with nephrolithiasis, and repressed apoptosis and the expression of OPN and CD44. Moreover, vitexin mitigated F4/80-positive macrophage infiltration and MCP-1 expression in the kidneys. Furthermore, an in vitro study showed that vitexin increased the viability of HK-2 cells and THP-1-derived macrophages, which was impaired by treatment with COM crystals, decreased the medium lactate dehydrogenase (LDH) level, and inhibited the expression of pyroptosis-related proteins in HK-2 cells and macrophages. Vitexin repressed EMT of HK-2 cells, with increased expression of pan-cytokeratin (Pan-ck) and decreased expression of Vimentin and alpha-smooth muscle actin (α-SMA), and downregulated the Wnt/ß-catenin pathway. Moreover, vitexin suppressed tumor necrosis factor-α (TNF-α) and IL-1ß mRNA expression, which was upregulated by COM in macrophages. CONCLUSION: Vitexin exerts protective effects against nephrolithiasis by inhibiting pyroptosis activation, apoptosis, EMT, and macrophage infiltration. In addition, GSDMD-related pyroptosis mediates nephrolithiasis.

Methanolic extract of Cucumis melo attenuates ethylene glycol-induced nephrolithiasis in Wistar rats.

Evaluation of the effects of methanolic extract of Cucumis melo in ethylene glycol-induced nephrolithiasis on Wistar rats. 0.75% solution of ethylene glycol (EG) in payable water was given to produce nephrolithiasis on Wistar rats. The action of oral intake of methanolic extract of Cucumis melo seed in nephrolithiasis is studied and is matched with the action of oral intake of Cystone (standard) on Wistar rats. EG resulted in hyperoxaluria and deposition of calcium oxalate as well as raised urinary excretion of oxalate and calcium. Supplementation with methanolic extract of Cucumis melo seed decreased the increased renal oxalate, indicating a regulatory effect on oxalate formation endogenously. The outcomes stipulate that the seed of Cucumis melo is endowed with antinephrolithiatic action.

Antiurolithic effects of medicinal plants: results of in vivo studies in rat models of calcium oxalate nephrolithiasis-a systematic review.

Urolithiasis is one of the oldest diseases affecting humans, while plants are one of our oldest companions providing food, shelter, and medicine. In spite of substantial progress in understanding the pathophysiological mechanisms, treatment options are still limited, often expensive for common people in most parts of the world. As a result, there is a great interest in herbal remedies for the treatment of urinary stone disease as an alternative or adjunct therapy. Numerous in vivo and in vitro studies have been carried out to understand the efficacy of herbs in reducing stone formation. We adopted PRISMA guidelines and systematically reviewed PubMed/Medline for the literature, reporting results of various herbal products on in vivo models of nephrolithiasis/urolithiasis. The Medical Subject Heading Terms (Mesh term) "Urolithiasis" was used with Boolean operator "AND" and other related Mesh Unique terms to search all the available records (July 2019). A total of 163 original articles on in vivo experiments were retrieved from PubMed indexed with the (MeshTerm) "Urolithiasis" AND "Complementary Therapies/Alternative Medicine, "Urolithiasis" AND "Plant Extracts" and "Urolithiasis" AND "Traditional Medicine". Most of the studies used ethylene glycol (EG) to induce hyperoxaluria and nephrolithiasis in rats. A variety of extraction methods including aqueous, alcoholic, hydro-alcoholic of various plant parts ranging from root bark to fruits and seeds, or a combination thereof, were utilized. All the investigations did not study all aspects of nephrolithiasis making it difficult to compare the efficacy of various treatments. Changes in the lithogenic factors and a reduction in calcium oxalate (CaOx) crystal deposition in the kidneys were, however, considered favorable outcomes of the various treatments. Less than 10% of the studies examined antioxidant and diuretic activities of the herbal treatments and concluded that their antiurolithic activities were a result of antioxidant, anti-inflammatory, and/or diuretic effects of the treatments.

Nephroprotective and antilithiatic activities of Costus spicatus (Jacq.) Sw.: Ethnopharmacological investigation of a species from the Dourados region, Mato Grosso do Sul State, Brazil.

ETHNOPHARMACOLOGICAL RELEVANCE: Costus spicatus (Jacq.) Sw., also known as "cana-do-brejo," is a species that is widely used in Brazilian traditional medicine for the treatment of kidney diseases. However, no studies have evaluated its nephroprotective and antilithiatic effects. AIM: To investigate nephroprotective and antilithiatic effects of C. spicatus in a preclinical model of acute kidney injury (AKI) and in vitro nephrolithiasis. MATERIALS AND METHODS: C. spicatus leaves were collected directly from the natural environment in the Dourados region, Mato Grosso do Sul State, Brazil. The ethanol-soluble fraction of C. spicatus (ESCS) was obtained by infusion. Phytochemical characterization was performed by liquid chromatography coupled to diode array detector and mass spectrometer (LC-DAD-MS). We assessed whether ESCS has acute or prolonged diuretic activity. The nephroprotective effects of ESCS were evaluated in a model of AKI that was induced by glycerol (10 ml/kg, intramuscularly) in Wistar rats. Different doses of ESCS (30, 100, and 300 mg/kg) were administered orally for 5 days before the induction of AKI. Urinary parameters were measured on days 1, 3, 5, and 7. Twenty-four hours after the last urine collection, blood samples were obtained for the biochemical analysis. Blood pressure levels, renal vascular reactivity, renal tissue redox status, and histopathological changes were measured. Antilithiatic effects were evaluated by in vitro crystallization. Calcium oxalate precipitation was induced by sodium oxalate in urine samples with ESCS at 0.05, 0.5, and 5 mg/ml. RESULTS: From LC-DAD-MS analyses, flavonoids, saponins and other phenolic compounds were determined in the composition of ESCS. Significant reductions of the excretion of urinary total protein, creatinine, sodium, and potassium were observed in the AKI group, with significant histopathological damage (swelling, vacuolization, necrosis, and inflammatory infiltration) in the proximal convoluted tubule. Treatment with ESCS exerted a significant nephroprotective effect by increasing the urinary excretion of total protein, urea, creatinine, sodium, potassium, calcium, and chloride. All of the groups that were treated with ESCS exhibited a reduction of histopathological lesions and significant modulation of the tissue redox state. We also observed a concentration-dependent effect of ESCS on the crystallization of urinary crystals, with reductions of the size and proportion of monohydrated crystals. CONCLUSION: The data suggest that C. spicatus has nephroprotective and antilithiatic effects, suggesting possible effectiveness in its traditional use.