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Item Collagen fibrils and cell nuclei are entrapped within Randall's plaques but not in CaOx matrix overgrowth: A microscopic inquiry into Randall's plaque stone pathogenesis(Wiley, 2022) Canela, Victor Hugo; Bledsoe, Sharon B.; Worcester, Elaine M.; Lingeman, James E.; El-Achkar, Tarek M.; Williams, James C., Jr.; Anatomy, Cell Biology and Physiology, School of MedicineCalcium oxalate (CaOx) stones can grow attached to the renal papillary calcification known as Randall's plaque. Although stone growth on Randall's plaque is a common phenomenon, this mechanism of stone formation is still poorly understood. The objective of this study was to investigate the microenvironment of mature Randall's plaque, explore its molecular composition and differentiate plaque from CaOx overgrowth using multimodal imaging on demineralized stone sections. Fluorescence imaging showed consistent differences in autofluorescence patterns between Randall's plaque and calcium oxalate overgrowth regions. Second harmonic generation imaging established the presence of collagen only in regions of decalcified Randall's plaque but not in regions of CaOx overgrowth matrix. Surprisingly, in these stone sections we observed cell nuclei with preserved morphology within regions of mature Randall's plaque. These conserved cells had variable expression of vimentin and CD45. The presence of nuclei in mature plaque indicates that mineralization is not necessarily associated with cell death. The markers identified suggest that some of the entrapped cells may be undergoing dedifferentiation or could emanate from a mesenchymal or immune origin. We propose that entrapped cells may play an important role in the growth and maintenance of Randall's plaque. Further characterization of these cells and thorough analyses of the mineralized stone forming renal papilla will be fundamental in understanding the pathogenesis of Randall's plaque and CaOx stone formation.Item Effects of DCPD Cement Chemistry on Degradation Properties and Cytocompatibility: Comparison of MCPM/β-TCP and MCPM/HA Formulations(IOP, 2013) Alge, Daniel L.; Goebel, W. Scott; Chu, Tien-Min Gabriel; Pediatrics, School of MedicineDicalcium phosphate dihydrate (DCPD) cements are attractive biomaterials for bone repair, and a number of different DCPD cement formulations have been proposed in the literature. In this study, we have specifically compared monocalcium phosphate monohydrate (MCPM)/hydroxyapatite (HA) and MCPM/β-tricalcium phosphate (β-TCP) formulations to test the hypothesis that DCPD cement chemistry affects the degradation properties and cytocompatibility of the cement. Using simple in vitro models we found that MCPM/β-TCP formulations degraded primarily by DCPD dissolution, which was associated with a slight pH drop and relatively low mass loss. Cytocompatibility testing of cement conditioned culture media revealed no significant change in cell viability relative to the negative control for all of the MCPM/β-TCP formulations. In contrast, the MCPM/HA formulations were prone to undergo rapid conversion of DCPD to HA, resulting in a sharp pH drop and extensive mass loss. A stoichiometric excess of HA in the cement was found to accelerate the conversion process, and significant cytotoxicity was observed for the MCPM/HA formulations containing excess HA. Collectively, these results show that, although the product of the setting reaction is the same, DCPD cements produced with MCPM/HA and MCPM/β-TCP formulations differ significantly in their degradation properties and cytocompatibility. These differences may have important implications for the selection of a DCPD cement formulation for clinical application.Item Phenotypic characterization of kidney stone formers by endoscopic and histological quantification of intra-renal calcification(Elsevier, 2013) Linnes, Michael P.; Krambeck, Amy E.; Cornell, Lynn; Williams, James C., Jr.; Korinek, Mark; Bergstralh, Eric J.; Li, Xujian; Rule, Andrew D.; McCollough, Cynthia M.; Vrtiska, Terri J.; Lieske, John; Anatomy, Cell Biology and Physiology, School of MedicineInterstitial Randall's plaques and collecting duct plugs are distinct forms of renal calcification thought to provide sites for stone retention within the kidney. Here we assessed kidney stone precursor lesions in a random cohort of stone formers undergoing percutaneous nephrolithotomy. Each accessible papilla was endoscopically mapped following stone removal. The percent papillary surface area covered by plaque and plug were digitally measured using image analysis software. Stone composition was determined by micro-computed tomography and infrared analysis. A representative papillary tip was biopsied. The 24-h urine collections were used to measure supersaturation and crystal growth inhibition. The vast majority (99%) of stone formers had Randall's plaque on at least 1 papilla, while significant tubular plugging (over 1% of surface area) was present in about one-fifth of patients. Among calcium oxalate stone formers the amount of Randall's plaque correlated with higher urinary citrate levels. Tubular plugging correlated positively with pH and brushite supersaturation but negatively with citrate excretion. Lower urinary crystal growth inhibition predicted the presence of tubular plugging but not plaque. Thus, tubular plugging may be more common than previously recognized among patients with all types of stones, including some with idiopathic calcium oxalate stones.Item Three-Dimensional Printing of Clinical Scale and Personalized Calcium Phosphate Scaffolds for Alveolar Bone Reconstruction(Elsevier, 2022) Anderson, Margaret; Dubey, Nileshkumar; Bogie, Kath; Cao, Chen; Li, Junying; Lerchbacker, Joseph; Mendonça, Gustavo; Kauffman, Frederic; Bottino, Marco C.; Kaigler, Darnell; Biomedical and Applied Sciences, School of DentistryObjective: Alveolar bone defects can be highly variable in their morphology and, as the defect size increases, they become more challenging to treat with currently available therapeutics and biomaterials. This investigation sought to devise a protocol for fabricating customized clinical scale and patient-specific, bioceramic scaffolds for reconstruction of large alveolar bone defects. Methods: Two types of calcium phosphate (CaP)-based bioceramic scaffolds (alginate/β-TCP and hydroxyapatite/α-TCP, hereafter referred to as hybrid CaP and Osteoink™, respectively) were designed, 3D printed, and their biocompatibility with alveolar bone marrow stem cells and mechanical properties were determined. Following scaffold optimization, a workflow was developed to use cone beam computed tomographic (CBCT) imaging to design and 3D print, defect-specific bioceramic scaffolds for clinical-scale bone defects. Results: Osteoink™ scaffolds had the highest compressive strength when compared to hybrid CaP with different infill orientation. In cell culture medium, hybrid CaP degradation resulted in decreased pH (6.3) and toxicity to stem cells; however, OsteoInk™ scaffolds maintained a stable pH (7.2) in culture and passed the ISO standard for cytotoxicity. Finally, a clinically feasible laboratory workflow was developed and evaluated using CBCT imaging to engineer customized and defect-specific CaP scaffolds using OsteoInk™. It was determined that printed scaffolds had a high degree of accuracy to fit the respective clinical defects for which they were designed (0.27 mm morphological deviation of printed scaffolds from digital design). Significance: From patient to patient, large alveolar bone defects are difficult to treat due to high variability in their complex morphologies and architecture. Our findings shows that Osteoink™ is a biocompatible material for 3D printing of clinically acceptable, patient-specific scaffolds with precision-fit for use in alveolar bone reconstructive procedures. Collectively, emerging digital technologies including CBCT imaging, 3D surgical planning, and (bio)printing can be integrated to address this unmet clinical challenge.Item Variability in stone composition and metabolic correlation between kidneys in patients with bilateral nephrolithiasis(Elsevier, 2019-12-20) Rivera, Marcelino E.; Nottingham, Charles U.; Borofsky, Michael S.; Kissel, Suzanne M.; Maniar, Viraj; Dauw, Casey A.; York, Nadya E.; Krambeck, Amy E.; Lingeman, James E.; Urology, School of MedicineIntroduction: To evaluate the clinical significance of discordant stone analyses in patients undergoing bilateral ureteroscopy. Methods: A retrospective chart review was performed for all patients undergoing stone extraction with bilateral ureteroscopy at our institution in an aim to identify patients who had bilateral stone analysis and 24-hour urine chemistry data available. Stones were then classified based upon the dominant present (>50%). Twenty-four hour urinalysis results were reviewed and statistical analysis performed comparing discordant and concordant patient populations, assessing significant differences that would potentially influence clinical management. Results: We identified 79 patients (158 renal units) who had bilateral stones removed at the time of ureteroscopy. The majority of stones were classified as calcium oxalate (CaOx) (60.1%) followed by calcium phosphate (CaP) (27.8%), brushite (5.1%), uric acid (UA) (4.4%) and cystine (2.5%). Discrepancies in stone classifications were present 24% of the time. Evaluation of 24-hour urinalysis results demonstrated that patients with CaOx:CaP stone discordance compared to CaOx:CaOx concordant stone formers were more likely to have an elevated pH (p=0.02) and lower uric acid supersaturation (p=0.01). Conclusions: Discrepancies in stone mineral content are common in patients with bilateral stone disease. A single stone analysis from one side in the setting of bilateral stone disease is insufficient for management of patients with bilateral renal stones, and may lead to mismanagement when this misrepresented information is utilized in addition to 24-hour urinalysis results. At least one stone analysis should be performed from both sides during a bilateral stone extraction procedure.