Calcium Oxalate Dihydrate Crystals Research Articles

Carboxymethylated Desmodium styracifolium polysaccharide reduces the risk of calcium oxalate kidney stone formation by inhibiting crystal adhesion and promoting crystal endocytosis.

The inhibition of cell surface crystal adhesion and an appropriate increase in crystal endocytosis contribute to the inhibition of kidney stone formation. In this study, we investigated the effects of different degrees of carboxymethylation on these processes. An injury model was established by treating human renal proximal tubular epithelial (HK-2) cells with 98.3 ± 8.1 nm calcium oxalate dihydrate (nanoCOD) crystals. The HK-2 cells were protected with carboxy (-COOH) Desmodium styracifolium polysaccharides at 1.17% (DSP0), 7.45% (CDSP1), 12.2% (CDSP2), and 17.7% (CDSP3). Changes in biochemical indexes and effects on nanoCOD adhesion and endocytosis were detected. The protection of HK-2 cells from nanoCOD-induced oxidative damage by carboxymethylated Desmodium styracifolium polysaccharides (CDSPs) is closely related to the protection of subcellular organelles, such as mitochondria. CDSPs can reduce crystal adhesion on the cell surface and maintain appropriate crystal endocytosis, thereby reducing the risk of kidney stone formation. CDSP2 with moderate -COOH content showed the strongest protective activity among the CDSPs.

The impact of crystal phase transition on the hardness and structure of kidney stones

Calcium oxalate kidney stones, the most prevalent type of kidney stones, undergo a multi-step process of crystal nucleation, growth, aggregation, and secondary transition. The secondary transition has been rather overlooked, and thus, the effects on the disease and the underlying mechanism remain unclear. Here, we show, by periodic micro-CT images of human kidney stones in an ex vivo incubation experiment, that the growth of porous aggregates of calcium oxalate dihydrate (COD) crystals triggers the hardening of the kidney stones that causes difficulty in lithotripsy of kidney stone disease in the secondary transition. This hardening was caused by the internal nucleation and growth of precise calcium oxalate monohydrate (COM) crystals from isolated urine in which the calcium oxalate concentrations decreased by the growth of COD in closed grain boundaries of COD aggregate kidney stones. Reducing the calcium oxalate concentrations in urine is regarded as a typical approach for avoiding the recurrence. However, our results revealed that the decrease of the concentrations in closed microenvironments conversely promotes the transition of the COD aggregates into hard COM aggregates. We anticipate that the suppression of the secondary transition has the potential to manage the deterioration of kidney stone disease.

Different Degrees of Sulfated Laminaria Polysaccharides Recovered Damaged HK-2 Cells and Inhibited Adhesion of Nano-COM and Nano-COD Crystals.

The crystal adhesion caused by the damage of renal tubular epithelial cells (HK-2) is the key to the formation of kidney stones. However, no effective preventive drug has been found. This study aims to explore the recovery effects of four Laminaria polysaccharides (SLPs) with different sulfate (-OSO3-) contents on damaged HK-2 cells and the difference in the adhesion of damaged cells to nanometer calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD) before and after recovery. Sodium oxalate (2.6 mmol/L) was used to damage HK-2 cells to establish a damaged model. SLPs (LP0, SLP1, SLP2, and SLP3) with -OSO3- contents of 0.73%, 15.1%, 22.8%, and 31.3%, respectively, were used to restore the damaged cells, and the effects of SLPs on the adhesion of COM and COD, with a size of about 100 nm before and after recovery, were measured. The following results were observed after SLPs recovered the damaged HK-2 cells: increased cell viability, restored cell morphology, decreased reactive oxygen levels, increased mitochondrial membrane potential, decreased phosphatidylserine eversion ratio, increased cell migration ability, reduced expression of annexin A1, transmembrane protein, and heat shock protein 90 on the cell surface, and reduced adhesion amount of cells to COM and COD. Under the same conditions, the adhesion ability of cells to COD crystals was weaker than that to COM crystals. As the sulfate content in SLPs increases, the ability of SLPs to recover damaged HK-2 cells and inhibit crystal adhesion increases. SLP3 with high -OSO3- content may be a potential drug to prevent kidney stones.

Carboxymethylated Rhizoma alismatis Polysaccharides Regulate Calcium Oxalate Crystals Growth and Reduce the Regulated Crystals' Cytotoxicity.

This study explored the effects of polysaccharides (RAPD) extracted from the traditional anti-stone Chinese medicine Rhizoma alismatis and their carboxymethylated derivatives (RAPs) on the crystal phase, morphology, and size of calcium oxalate (CaOx). It also determined the damaging ability of the regulated crystals on human renal tubular epithelial cells (HK-2). RAPD carboxymethylation with a carboxyl group (-COOH) content of 3.57% was carried out by the chloroacetic acid solvent method. The effects of -COOH content in RAPs and RAP concentration on the regulation of CaOx crystal growth were studied by controlling the variables. Cell experiments were conducted to explore the differences in the cytotoxicity of RAP-regulated crystals. The -COOH contents of RAPD, RAP1, RAP2, and RAP3 were 3.57%, 7.79%, 10.84%, and 15.33%, respectively. RAPs can inhibit the growth of calcium oxalate monohydrate (COM) and induce the formation of calcium oxalate dihydrate (COD). When the -COOH content in RAPs was high, their ability to induce COD formation was enhanced. In the crystals induced by RAPs, a high COD content can lower the damage to cells. In particular, the cytotoxicity of the crystals induced by RAP3 was the lowest. When the concentration of RAP3 increased, the cytotoxicity gradually increased due to the reduced size of the formed COD crystals. An interaction was observed between RAPs and crystals, and the number of RAPs adsorbed in the crystals was positively correlated with the -COOH content in RAPs. RAPs can reduce the damage of CaOx to HK-2 cells by regulating the crystallization of CaOx crystals and effectively reducing the risk of kidney stone formation. RAPs, especially RAP3 with a high carboxyl group content, has the potential to be developed as a novel green anti-stone drug.

MP05-16 THE DEVELOPMENT OF A THREE-DIMENSIONAL KIDNEY SPHEROID MODEL TO STUDY CALCIUM-BASED CRYSTAL FORMATION

You have accessJournal of UrologyCME1 Apr 2023MP05-16 THE DEVELOPMENT OF A THREE-DIMENSIONAL KIDNEY SPHEROID MODEL TO STUDY CALCIUM-BASED CRYSTAL FORMATION John Chmiel, Wongsakorn Kiattiburut, Aaron Kwong, Priyanka Prabhu, Hassan Razvi, Leonard Luyt, Jeremy Burton, and Jennifer Bjazevic John ChmielJohn Chmiel More articles by this author , Wongsakorn KiattiburutWongsakorn Kiattiburut More articles by this author , Aaron KwongAaron Kwong More articles by this author , Priyanka PrabhuPriyanka Prabhu More articles by this author , Hassan RazviHassan Razvi More articles by this author , Leonard LuytLeonard Luyt More articles by this author , Jeremy BurtonJeremy Burton More articles by this author , and Jennifer BjazevicJennifer Bjazevic More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003216.16AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: An estimated 10% of North Americans suffer from nephrolithiasis. With rising incidence rates, there is a strong need for models to study the pathophysiology of stone disease. In vitro cell culture models are essential for mechanistic and high-throughput analysis of stone formation and drug treatment; however, there are no suitable models that adequately represent the pathophysiology of stone disease. Advancements in cell culture have given rise to three-dimensional cysts, which are hollow spheroid structures that have apical to basal polarization. These culture systems have similar physiochemical environments to in vivo models and can be cultured for the extended times required for stone formation. Here, we developed a novel 3D kidney cyst model that can generate calcium-based kidney stones. METHODS: Madin-Darby canine kidney (MDCK) cells were cultured in suspension in growth factor-reduced Matrigel to generate hollow spheroid cysts. Crystal formation was induced by culturing cysts in a custom crystal-forming media for two weeks. Immunofluorescent staining was used for characterizing and visualizing the three-dimensional structure. Alendronate-fluorescein, a fluorescently labelled bisphosphonate that specifically binds to calcium-containing crystals, was used along with plain polarized light to visualize the crystal structures. RESULTS: Stains for cellular components and structures demonstrated that the luminal spheroids were successfully formed under the culture conditions. The spheroids also have basal-to-apical polarization suggesting that crystals will form on the inside lumen of the hollow cyst. Birefringent structures resembling calcium oxalate dihydrate and calcium phosphate crystals were observed embedded along the apical face of the spheroid and protruding into the lumen. Alendronate-fluorescein staining confirmed the presence of calcium-containing crystals in the lumen of the cyst. CONCLUSIONS: We have demonstrated that hollow MDCK spheroids cultured with crystal-forming media can generate calcium-containing crystals in the spheroid lumen. This model is the first in vitro tissue culture-based model to reliably develop calcium-based crystals and can be used to investigate their pathogenesis or to perform high-throughput drug screening. Furthermore, it paves the way for personalized medicine by building the foundation for a cell system where renal tissue can be harvested and cultured to identify individual intrinsic risk factors that might contribute to stone development. Source of Funding: Canadian Institutes of Health Research (CIHR) CGS-D, The Northeastern Section of the American Urological Association (NSAUA), Lawson Health Research © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e49 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information John Chmiel More articles by this author Wongsakorn Kiattiburut More articles by this author Aaron Kwong More articles by this author Priyanka Prabhu More articles by this author Hassan Razvi More articles by this author Leonard Luyt More articles by this author Jeremy Burton More articles by this author Jennifer Bjazevic More articles by this author Expand All Advertisement PDF downloadLoading ...

MP05-20 KIDNEY STONE GROWTH ‘RINGS’ VISUALIZED USING MICRO CT AND FLUORESCENCE MICROSCOPY: SIMILARITIES IN GROWTH OF RANDALL’S PLAQUE CALCIUM OXALATE STONES FROM THE SAME KIDNEYS

You have accessJournal of UrologyCME1 Apr 2023MP05-20 KIDNEY STONE GROWTH ‘RINGS’ VISUALIZED USING MICRO CT AND FLUORESCENCE MICROSCOPY: SIMILARITIES IN GROWTH OF RANDALL’S PLAQUE CALCIUM OXALATE STONES FROM THE SAME KIDNEYS Emily Wachter, Angela Sabo, Tarek El-Achkar, Elaine Worcester, James Lingeman, and James Williams Emily WachterEmily Wachter More articles by this author , Angela SaboAngela Sabo More articles by this author , Tarek El-AchkarTarek El-Achkar More articles by this author , Elaine WorcesterElaine Worcester More articles by this author , James LingemanJames Lingeman More articles by this author , and James WilliamsJames Williams More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003216.20AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Recent work has shown that mechanisms of stone growth can be demonstrated at the microscopic level using fluorescence microscopy. The objective here was to study multiple stones to observe patterns of mineral and organic deposition within and between patients. METHODS: Stones were removed by basket during endoscopic procedure, and a total of 12 stones were studied from 3 patients. All stones were verified by micro CT as having grown on Randall’s plaque by residue visible on the stone. Each stone was mounted on polystyrene and ground down to reveal a planar surface inside the stone. The stone was then imaged using confocal microscopy (Leica SP8) with a water immersion lens (20x, 0.75 NA). Each stone was ground repeatedly to collect as many interior planes as possible, with micro CT verification of each plane of section and mineral regions exposed. RESULTS: Within a patient, similarities of layering (both thickness and color) were apparent in stones from the same kidney, but stones from different patients had different fluorescent features. Several stones showed evidence of episodic growth by deposition of aggregates of calcium oxalate dihydrate crystals, and these stones with evidence of dihydrate aggregates tended to be larger stones within a given patient. Regions of apatite adjacent to Randall’s plaque showed yellow fluorescence distinctly different from apatite in later stone overgrowth. CONCLUSIONS: The unique fluorescence of apatite laid down as the first overgrowth on Randall’s plaque is suggestive of special urine molecules deposited at the initiation of stone growth. Outside of this consistent finding, fluorescent molecules laid down with mineral in calcium oxalate stones are not universal among patients, but stones from the same kidney showed similar fluorescence patterns. Source of Funding: NIH R01 DK124776, NIH P01 DK056788 © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e52 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Emily Wachter More articles by this author Angela Sabo More articles by this author Tarek El-Achkar More articles by this author Elaine Worcester More articles by this author James Lingeman More articles by this author James Williams More articles by this author Expand All Advertisement PDF downloadLoading ...

Detection and identification of urinary crystals using an automated urine particle analyzer coupled with a digital particle imaging device

Polarized light microscopy (POM) remains the gold standard for crystalluria analysis. However, such method is time consuming and requires well-trained staff. Here, to address this issue, we tested the Sysmex UF-4000 analyzer coupled to a UD10 module as an automated flow cytometry-digital particle imaging workflow to assess (i) the ability of the system to detect and identify the crystals species and (ii) the quality of the images provided by the UD-10 module (n = 40) for each urine sample analyzed. First, systematic analysis of 76 samples by POM and the UF-4000/UD-10 analyzer showed that only attentive examination of the 40 photos was able to confidently detect crystalluria-positive samples with no misses and thus serve to discriminate positive-test crystalluria from negative-test crystalluria. These first results were confirmed by sensitivity analysis and the negative predictive value calculated on 200 samples for the results provided by the UF-4000 (39% and 46%) and after examination of the 40 UD-10 photos (100% for the both values). Digital images can therefore serve to screen crystalluria without missing crystals. A part of samples were treated by POM whereas it was not necessary (positive predictive value: 78%). Finally, we compared the crystal identification performances of the Sysmex UF4000/UD10 workflow and the ‘gold standard’ POM method on 131 urine samples containing crystals. Only calcium oxalate dihydrate crystals were identified by the Sysmex UF-4000. A close examination of the digital photographs enabled exact identification of crystals in 84.7% of the samples, suggesting however that POM is still require as soon as crystals are observed on the photographs. We conclude that a SYSMEX UF-4000 coupled with a UD-10 module can be used in practice with close examination of the photographs to discriminate positive crystalluria from negative crystalluria.

Multicellular spheroids containing synthetic mineral particles: an advanced 3D tumor model system to investigate breast precancer malignancy potential according to the mineral type.

Mineral particles that form in soft tissues in association with disease conditions are heterogeneous in their composition and physiochemical properties. Hence, it is challenging to study the effect of mineral type on disease progression in a high-throughput and realistic manner. For example, most early breast precancer lesions, termed ductal carcinoma in situ (DCIS), contain microcalcifications (MCs), calcium-containing pathological minerals. The most common type of MCs is calcium phosphate crystals, mainly carbonated apatite; it is associated with either benign or malignant lesions. In vitro studies indicate that the crystal properties of apatite MCs can affect breast cancer progression. A less common type of MCs is calcium oxalate dihydrate (COD), which is almost always found in benign lesions. We developed a 3D tumor model of multicellular spheroids of human precancer cells containing synthetic MC analogs that link the crystal properties of MCs with the progression of breast precancer to invasive cancer. Using this 3D model, we show that apatite crystals induce Her2 overexpression in DCIS cells. This tumor-triggering effect is increased when the carbonate fraction in the MCs decreases. COD crystals, in contrast, decrease Her2 expression in the spheroids, even compared with a control group with no added MC analogs. Furthermore, COD decreases cell proliferation and migration in DCIS monolayers compared to untreated cells and cells incubated with apatite crystals. This finding suggests that COD is not randomly located only in benign lesions-it may actively contribute to suppressing precancer progression in its surroundings. Our model provides an easy-to-manipulate platform to better understand the interactions between mineral particles and their biological microenvironment. A better understanding of the effect of the crystal properties of MCs on precancer progression will potentially provide new directions for better precancer prognosis and treatment.

Effect of Feed Supplementation with Tripotassium Citrate or Sodium Chloride on the Development of Urinary Calcium Oxalate Crystals in Fattening Pigs.

The present study investigated whether dietary supplementation of tripotassium citrate or NaCl reduced the prevalence of calcium oxalate dihydrate (COD) uroliths in fattening pigs on a farm with a high prevalence of COD uroliths. Each group (control, TPC, NaCl) consisted of three batches of approximately 260 fattening pigs each. Performance, water intake, markers for bone resorption (CTX) and bone formation (osteocalcin) and urinalysis from samples taken at the farm and in the slaughterhouse were investigated. Performance parameters, feed and water intake, CTX and osteocalcin were not significantly different between the groups (p > 0.05). The main crystals found were struvite, COD, calcite and amorphous crystals. The prevalence of COD crystals was lower in samples from the slaughterhouse in each group. Microscopic and biochemical examination of urine showed large differences between samples from the farm and the slaughterhouse. In conclusion, there were no beneficial effects of feed supplementation with TPC or NaCl on the prevalence of COD crystals but TPC has a clear promoting influence on the development of alkaline calcite crystals. Urinalysis from samples taken at the slaughterhouse does not fully reflect the situation on the farm.

CLINICOPATHOLOGIC EVALUATION OF AN EX SITU POPULATION OF ADULT AFRICAN WHITE-BELLIED PANGOLIN (PHATAGINUS TRICUSPIS).

Death due to illegal trade and trafficking serves as the largest threat to pangolins in their geographic range countries. Appropriate veterinary care is essential to the welfare of sick or debilitated pangolins. Given the challenge of comprehensive physical examination of this species due to its unique anatomic features and poorly studied physiology, laboratory diagnostics can provide essential information to guide appropriate medical care. This retrospective study included previously analyzed blood and urine samples collected from adult (n = 17) African white-bellied pangolins (Phataginus tricuspis) under managed care that were sampled over a 4-yr period as part of routine veterinary examinations. Samples from clinically normal animals were used to summarize results for this species. Pangolins had a lymphocyte-predominant leukogram with a mean neutrophil to lymphocyte ratio of 0.83 (min-max: 0.13-4.81). Seasonal, sex, and age differences were present for several hematologic and serum biochemical analytes. Urine samples had a median urine specific gravity (USG) of 1.040 (min-max: 1.006-1.060), a neutral-to-basic urine pH (6.0-8.6), and contained mainly struvite crystals or infrequently calcium oxalate dihydrate crystals. Winter samples had higher USG than summer samples. The clinicopathologic data provided herein offers applicable information useful for the clinical management of pangolins.

In Situ Identification of Unknown Crystals in Acute Kidney Injury Using Raman Spectroscopy.

Raman spectroscopy is a well-established and powerful tool for in situ biomolecular evaluation. Type 2 crystal nephropathies are characterized by the deposition of crystalline materials in the tubular lumen, resulting in rapid onset of acute kidney injury without specific symptoms. Timely crystal identification is essential for its diagnosis, mechanism exploration and therapy, but remains challenging. This study aims to develop a Raman spectroscopy-based method to assist pathological diagnosis of type 2 crystal nephropathies. Unknown crystals in renal tissue slides from a victim suffered extensive burn injury were detected by Raman spectroscopy, and the inclusion of crystals was determined by comparing Raman data with established database. Multiple crystals were scanned to verify the reproducibility of crystal in situ. Raman data of 20 random crystals were obtained, and the distribution and uniformity of substances in crystals were investigated by Raman imaging. A mouse model was established to mimic the crystal nephropathy to verify the availability of Raman spectroscopy in frozen biopsy. All crystals on the human slides were identified to be calcium oxalate dihydrate, and the distribution and content of calcium oxalate dihydrate on a single crystal were uneven. Raman spectroscopy was further validated to be available in identification of calcium oxalate dihydrate crystals in the biopsy specimens. Here, a Raman spectroscopy-based method for in situ identification of unknown crystals in both paraffin-embedded tissues and biopsy specimens was established, providing an effective and promising method to analyze unknown crystals in tissues and assist the precise pathological diagnosis in both clinical and forensic medicine.

Formation and Characterization of Irreversible Sediment of Ginseng Extract.

Sediment is a key issue in the beverage industry. This study confirmed that reversible and irreversible sediments were formed during low-temperature storage of ginseng extract. The first 30 days of storage are the critical period for sediment formation. As the time of storage extends, the chemical composition changes. The composition interaction model verified that the cross-linking of protein–pectin, protein–oxalic acid and Ca2+–pectin was the main cause of the turbidity of ginseng extract. Based on the characterization of irreversible sediment (IRS), there are typical structures of proteins, polysaccharides and calcium oxalate dihydrate (COD) crystals. Glucose, galacturonic acid, aspartate, glutamic acid, leucine, Ca, K, Al, Mg, Na and Fe are the main monomer components. Effective regulation of these ingredients will greatly help the quality of ginseng beverages.

Multicolor imaging of calcium-binding proteins in human kidney stones for elucidating the effects of proteins on crystal growth

The pathogenesis of kidney stone formation includes multi-step processes involving complex interactions between mineral components and protein matrix. Calcium-binding proteins in kidney stones have great influences on the stone formation. The spatial distributions of these proteins in kidney stones are essential for evaluating the in vivo effects of proteins on the stone formation, although the actual distribution of these proteins is still unclear. We reveal micro-scale distributions of three different proteins, namely osteopontin (OPN), renal prothrombin fragment 1 (RPTF-1), and calgranulin A (Cal-A), in human kidney stones retaining original mineral phases and textures: calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD). OPN and RPTF-1 were distributed inside of both COM and COD crystals, whereas Cal-A was distributed outside of crystals. OPN and RPTF-1 showed homogeneous distributions in COM crystals with mosaic texture, and periodically distributions parallel to specific crystal faces in COD crystals. The unique distributions of these proteins enable us to interpret the different in vivo effects of each protein on CaOx crystal growth based on their physico-chemical properties and the complex physical environment changes of each protein. This method will further allow us to elucidate in vivo effects of different proteins on kidney stone formation.

Effect of L-alanyl-glycine dipeptide on calcium oxalate crystallization in artificial urine

Pathological crystallization of calcium oxalate (CaOx), the most common constituent of kidney stones, has attracted much attention due to recent surge in reported natural and synthetic additives effectively inhibiting its nucleation and growth. The aim of this study is to investigate the effect of L-alanyl–glycine (Ala–Gly), a dipeptide commonly found in human urine, on CaOx crystallization and its phase transformation in the presence of an artificial urine media. The nucleated CaOx crystals are characterized by XRD, FTIR, SEM, and dynamic light scattering in terms of changes in their crystalline form, morphology, and size. XRD and FTIR results revealed that Ala–Gly inhibited the formation of the thermodynamically most stable phase of CaOx, calcium oxalate monohydrate (COM) crystals. SEM images revealed that hexagonal plate-shaped COM crystals are transformed into the smaller tetragonal bipyramidal calcium oxalate dihydrate (COD) crystals with increasing additive concentrations. At 125 ppm Ala–Gly concentration more pronounced aggregation of CaOx crystals is observed accompanied with higher negative zeta potential value of −27.1 ± 2.9 mV. Moreover, the phase transformation from COM to COD is also confirmed through thermogravimetric analysis. Consequently, these results suggest that Ala–Gly has a profound effect on preventing the formation of COM crystals and helping to stabilize the COD crystals, a CaOx phase that is reported to have a lower tendency to stick to kidney cells thus decreasing the risk of stone formation. The reported suppression of COM in the presence of Ala–Gly might be significant to clinicians in their attempt to develop a long-term effective treatment for kidney stones.

Inhibition of Calcium Oxalate Formation and Antioxidant Activity of Carboxymethylated Poria cocos Polysaccharides.

Three carboxymethylated Poria cocos polysaccharides (PCP-C1, PCP-C2, and PCP-C3) with -COOH contents of 6.13%, 10.24%, and 16.22%, respectively, were obtained by carboxymethylation of the original polysaccharide (PCP-C0), which has a molecular weight of 4 kDa and a carboxyl (-COOH) content of 2.54%. The structure of the PCP-Cs was characterized by FT-IR, 1H NMR, and 13C NMR spectra. The four PCP-Cs exhibited antioxidant activity, and their ability to scavenge radicals (hydroxyl and DPPH) and chelate ferrous ions was positively correlated with the degree of carboxymethylation. As the content of -COOH groups in the PCP-Cs increases, their ability to regulate the growth of calcium oxalate (CaOx) crystals was enhanced, thus inhibiting the growth of calcium oxalate monohydrate (COM) crystals and inducing the formation of more calcium oxalate dihydrate (COD) crystals. The formed CaOx crystal was more round and blunt, the absolute value of the Zeta potential on the crystal surface increased, and the aggregation between crystals was inhibited. Thermogravimetric analysis curves showed that the proportions of PCP-C0, PCP-C1, PCP-C2, and PCP-C3 incorporated into the crystal were 20.52%, 15.60%, 10.65%, and 9.78%, respectively, in the presence of 0.4 g/L PCP-Cs. PCP-C protection resisted oxidative damages of human kidney proximal tubular epithelial cells (HK-2) caused by oxalate, resulting in increased cell viability and superoxide dismutase activity and decreased reactive oxygen species levels, malondialdehyde content, and 8-hydroxy-deoxyguanosine expression. Hence, PCP-Cs, especially PCP-C3, can inhibit the formation of CaOx crystals and may have the potential to be an alternative antistone drug.

Effects of Selenized Astragalus Polysaccharide on the Adhesion and Endocytosis of Nanocalcium Oxalate Dihydrate after the Repair of Damaged HK-2 Cells.

An oxidative damage model of human proximal renal epithelial cells (HK-2) was established using oxalate damage. The repair effects of Astragalus polysaccharide (APS) and selenized APS (Se-APS) on damaged HK-2 cells were investigated. Differences in the adhesion and endocytosis of HK-2 cells to calcium oxalate dihydrate crystals with a size of approximately 100 nm before and after APS and Se-APS repair were also explored. The results showed that after being repaired by APS and Se-APS, HK-2 cells exhibited increased cell viability, restored cell morphology, reduced reactive oxygen species level, increased mitochondrial membrane potential, reduced phosphatidylserine eversion, and osteopontin expression. Moreover, the amount of adherent crystals on the cell surface decreased, but the amount of endocytic crystals increased. At the same concentration, Se-APS exhibited better repair effects on the damaged HK-2 cells than APS. All these findings revealed that Se-APS may be a potential drug candidate for inhibiting the formation of kidney stones.

Assessment of Paronychia Argentea Extraction on Kidney Stone by Using Calcium Oxalate Method

Urinary calculi are stones (urolithiasis) that can form anywhere in urinary tract outside of the kidneys and mostly composed of calcium oxalate and phosphate, additionally with elevated throughout the last two decades in the world. Chemical composition plays a major part in nephrolithiasis. Therefore, the high concentrations of lithogenic substances in urine enhance the crystallization method in urine tract system. The most kidney stones form from calcium oxalate, the present study was inspected the effect of the crude aqueous extract as well as the fractionated methanol extract (ethyl acetate, isopropanol, acetone and methanol residue) of paronychia argentea on the crystallization of calcium oxalate salts. The effect of aqueous extract and fractionated methanol extract on the size, number, type of calcium oxalate crystals. Paronychia argentea both the crude aqueous and the fractionated extract, especially ethyl acetate fraction have antiurolithic activity via reducing crystal size as well as activate the formation of calcium oxalate dihydrate (COD) crystals out from calcium oxalate monohydrate (COM) with increasing concentration of extract. The shifting of crystallization process to producing calcium oxalate dihydrate (COD) rather than oxalate monohydrate (COM) and the reducing the crystal size and calcium ion concentration, in addition to the diuretic action of extract plays an important role in controlling urolithiasis.

Antioxidant Sulfated Polysaccharide from Edible Red Seaweed Gracilaria birdiae is an Inhibitor of Calcium Oxalate Crystal Formation.

The genus Gracilaria synthesizes sulfated polysaccharides (SPs). Many of these SPs, including those synthesized by the edible seaweed Gracilaria birdiae, have not yet been adequately investigated for their use as potential pharmaceutical compounds. Previous studies have demonstrated the immunomodulatory effects of sulfated galactans from G. birdiae. In this study, a galactan (GB) was extracted from G. birdiae and evaluated by cell proliferation and antioxidant tests. GB showed no radical hydroxyl (OH) and superoxide (O2−) scavenging ability. However, GB was able to donate electrons in two further different assays and presented iron- and copper-chelating activity. Urolithiasis affects approximately 10% of the world’s population and is strongly associated with calcium oxalate (CaOx) crystals. No efficient compound is currently available for the treatment of this disease. GB appeared to interact with and stabilize calcium oxalate dihydrate crystals, leading to the modification of their morphology, size, and surface charge. These crystals then acquired the same characteristics as those found in healthy individuals. In addition, GB showed no cytotoxic effect against human kidney cells (HEK-293). Taken together, our current findings highlight the potential application of GB as an antiurolithic agent.

A facile preparation of rutin nanoparticles and its effects on controlled growth and morphology of calcium oxalate crystals

Switching of the growth-mode of calcium-containing crystals are of great research interest in materials science for the inhibition of pathological biomineralization. Rutin, a flavonol glycoside, was reported to have many pharmacological activities. However, the efficiency of rutin for practical therapeutic applications is still a challenge due to its low aqueous solubility and limited bioavailability. Herein, we report a facile method for preparation and characterization of rutin nanoparticles of ~200 nm in size and zeta potential of −22.70 ± 2.51 mV, based on anti-solvent co-precipitation using the synergistic effect of surfactants. Morphological and structural characterization of the nanoparticles was carried out with scanning electron microscopy (SEM), transmission electron microscopy (TEM), dynamic light scattering (DLS), Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), thermal gravimetric analysis (TGA), and powder X-ray diffraction (XRD) patterns. These results showed that rutin nanoparticles had a smaller particle size than raw rutin, and changed into an amorphous structure from the crystal structure of raw rutin. In vitro release profiles showed a sustained release rate of hydrophilic rutin nanoparticles corresponding to 83.5 ± 8.4% for a time period of 8 days. Furthermore, the influence of the rutin nanoparticles on the tuning of the growth and morphology of calcium oxalate (CaC2O4) crystals have been assessed using SEM, XRD patterns, TGA analysis and adsorption isotherms. The experimental results showed that the rutin nanoparticles could induce the formation of oval and octahedral calcium oxalate dihydrate (COD) crystals in a concentration-dependent manner. It is believed that the hydrogen bonding between the phenolic groups in rutin with carboxylate ions in crystals contributes to their increased adsorption and directing calcium oxalate monohydrate to dihydrate forms. These results indicated the potential application of rutin nanoparticles for in vivo lithiatic studies as well as for other pharmacological properties.

Modulation of Calcium Oxalate Crystal Growth and Protection from Oxidatively Damaged Renal Epithelial Cells of Corn Silk Polysaccharides with Different Molecular Weights.

Corn silk polysaccharide (CSP0; molecular weight = 124 kDa) was degraded by ultrasonication to obtain five degraded polysaccharides, namely, CSP1, CSP2, CSP3, CSP4, and CSP5, with molecular weights of 26.1, 12.2, 6.0, 3.5, and 2.0 kDa, respectively. The structures of these polysaccharides were characterized by FT-IR, 1H NMR, and 13C NMR analyses. The antioxidant activities, including scavenging ability for hydroxyl radicals and DPPH free radicals, chelation ability for Fe2+ ions, and reducing ability of CSP increased with decreased molecular weight of CSPs within 6.0 to 124 kDa. However, antioxidant activity weakened when the molecular weight of CSPs reached 3.5 and 2 kDa. CSP3 with a molecular weight of 6.0 kDa exhibited the strongest antioxidant activity. After protection with 60 μg/mL CSPs, the viability of human renal proximal tubular epithelial cells (HK-2) damaged by nano-COM crystals increased, the level of reactive oxygen species decreased, and the amount of COM crystal adhered onto the cell surface decreased. The ability of CSPs to protect cells from CaOx crystal damage was consistent with their antioxidant activity. CSPs can specifically combine with CaOx crystal to inhibit the conversion of calcium oxalate dihydrate crystal to calcium oxalate monohydrate crystal. All these results showed that the activity of CSPs was closely correlated with molecular weight. A very high or low molecular weight of CSPs was not conducive to their activity. CSPs, especially CSP3 with a molecular weight of 6.0 kDa, can be used as a potential antistone drug.