Recent advances on the mechanisms of kidney stone formation (Review).

Kidney stone disease is one of the oldest diseases known to medicine; however, the mechanisms of stone formation and development remain largely unclear. Over the past decades, a variety of theories and strategies have been developed and utilized in the surgical management of kidney stones, as a result of recent technological advances. Observations from the authors and other research groups suggest that there are five entirely different main mechanisms for kidney stone formation. Urinary supersaturation and crystallization are the driving force for intrarenal crystal precipitation. Randall's plaques are recognized as the origin of calcium oxalate stone formation. Sex hormones may be key players in the development of nephrolithiasis and may thus be potential targets for new drugs to suppress kidney stone formation. The microbiome, including urease­producing bacteria, nanobacteria and intestinal microbiota, is likely to have a profound effect on urological health, both positive and negative, owing to its metabolic output and other contributions. Lastly, the immune response, and particularly macrophage differentiation, play crucial roles in renal calcium oxalate crystal formation. In the present study, the current knowledge for each of these five aspects of kidney stone formation is reviewed. This knowledge may be used to explore novel research opportunities and improve the understanding of the initiation and development of kidney stones for urologists, nephrologists and primary care.

Lithogenic Potential of Ureaplasma in Chronic Prostatitis.

INTRODUCTION: The role of Ureaplasma spp. (UPs) in the pathogenesis of chronic prostatitis is debated. The lithogenic potential of UPs could be a risk factor for the development of chronic prostatitis. METHODS: A total of 143 patients with identification of UPs were retrospectively selected from a database including patients with prostatitis-like symptoms who were studied according to the same protocol including clinical, microbiological and microscopic evaluation, and transrectal prostate ultrasound. A control group of patients with negative UPs was considered including 393 with chronic bacterial prostatitis (CBP), 42 patients with Chlamydia trachomatis (CT), and 781 patients with chronic pelvic pain syndrome. UPs and Mycoplasma hominis (MH) were identified using a semiquantitative assay. RESULTS: Calcifications were observed more frequently in patients with UPs (64%) than in patients with CBP without UPs (39%), CT infection (37%), and chronic pelvic pain syndrome (29%) (p < 0.0001). UPs were isolated in VB1 alone in 35 patients (urethral UPs), in expressed prostatic secretion (EPS) or post-massage urine (VB3) or sperm in 77 patients (prostatic UPs) and associated with other pathogens in 31 patients (associated UPs). Calcifications were more frequent in prostatic UPs (71%) and associated UPs (73%) than in urethral UPs (34%). Mean NIH-CPSI scores were not significantly different between groups, although mean WBC counts of sperm of patients with urethral UPs were significantly lower than in patients with prostatic UPs (p = 0.000) and associated UPs (p = 0.002). CONCLUSIONS: UPs identification in the urogenital fluids is related to higher rates of prostate calcifications. The ability of UPs to promote the formation of calcifications could be related to the chronicization of prostate infection. In particular, the presence of UPs in VB3/EPS/sperm is associated with higher rates of calcifications and high WBC sperm counts, suggesting a partial or full causative role of UPs in the pathogenesis of this disease.

Biliary microbiota and mucin 4 impact the calcification of cholesterol gallstones.

BACKGROUND: Cholesterol gallstones account for over 80% of gallstones, and the pathogenesis of gallstone formation involves genetic and environmental factors. However, data on the evolution of cholesterol gallstones with various densities are limited. This study aimed to determine the roles of microbiota and mucins on the formation of calcified cholesterol gallstones in patients with cholelithiasis. METHODS: Paired gallbladder tissues and bile specimens were obtained from cholelithiasis patients who were categorized into the isodense group and calcified group according to the density of gallstones. The relative abundance of microbiota in gallbladder tissues was detected. Immunohistochemistry and enzyme-linked immunosorbent assay were performed to detect the expression levels of MUC1, MUC2, MUC3a, MUC3b, MUC4, MUC5ac and MUC5b in gallbladder tissues and bile. The correlation of microbiota abundance with MUC4 expression was evaluated by linear regression. RESULTS: A total of 23 patients with gallbladder stones were included. The density of gallstones in the isodense group was significantly lower than that of the calcified group (34.20 ± 1.50 vs. 109.40 ± 3.84 HU, P < 0.0001). Compared to the isodense group, the calcified group showed a higher abundance of gram-positive bacteria at the fundus, in the body and neck of gallbladder tissues. The concentrations of MUC1, MUC2, MUC3a, MUC3b, MUC5ac and MUC5b in the epithelial cells of gallbladder tissues showed no difference between the two groups, while the concentrations of MUC4 were significantly higher in the calcified group than that in the isodense group at the fundus (15.49 ± 0.69 vs. 10.23 ± 0.54 ng/mL, P < 0.05), in the body (14.54 ± 0.94 vs. 11.87 ± 0.85 ng/mL, P < 0.05) as well as in the neck (14.77 ± 1.04 vs. 10.85 ± 0.72 ng/mL, P < 0.05) of gallbladder tissues. Moreover, the abundance of bacteria was positively correlated with the expression of MUC4 (r = 0.569, P < 0.05) in the calcified group. CONCLUSIONS: This study showed the potential clinical relevance among biliary microbiota, mucins and calcified gallstones in patients with gallstones. Gram-positive microbiota and MUC4 may be positively associated with the calcification of cholesterol gallstones.

Quantification of Multiple Bacteria in Calcified Structural Valvular Heart Disease.

Genome studies of heart valve tissue (HVT) in patients with structural valvular heart disease (sVHD) and acute infective endocarditis (aIE) showed polymicrobial infections. Subject of this study is the quantification of bacterial DNA in HVT of sVHD in comparison to aIE. It will be examined whether the bacterial DNA concentration can be used as surrogate marker to differentiate chronic and acute infections. DNA was isolated from HVT of 100 patients with sVHD and 23 microbiologically positively tested patients with aIE. Selected pathogens (Cutibacterium acnes, Enterococcus faecalis, Enterococcus faecium, Staphylococcus aureus, Streptococcus pyogenes, Streptococcus agalactiae, Clostridium difficile, and Klebsiella pneumoniae) were quantified using TaqMan-qPCR. Polymicrobial infiltration of HVT was investigated by immunohistologic methods. Of 100 sVHD patients, 94 tested positive for bacteria by 16S-rDNA and 72 by TaqMan-qPCR. In 29% of the sVHD cohort and in 70% of the aIE cohort, a coinfection with more than 2 bacteria was observed as indication of a polymicrobial infection. The most common pathogens in the sVHD patients were C. acnes (59%; 5-4074 pg/mL), E. faecalis (16%, 174-2781 pg/mL), and S. aureus (15%, 8-105 pg/mL). The DNA concentration of E. faecalis (P = 0.0285) and S. aureus (P = 0.0149) is significantly lower in the sVHD cohort than in the aIE cohort. sVHD is associated with bacterial infection and infiltration of the HVT in a majority of cases. TaqMan-qPCR is a valid instrument for the specific detection of bacteria in HVT and allows discrimination between sVHD and aIE for E. faecalis and S. aureus.

Delayed septic radiation-induced calcinosis cutis long after cured anorectal cancer.

PURPOSE: Calcinosis cutis is an anecdotal local injury seen long after irradiation in cancer survivors. Our purpose was to shed light on this little studied and potentially serious ulceration. CASES: We report two cases of severe perineal-sacral infection with hard lesions, one decade after anorectal cancer irradiation. CT-scans showed extensive calcification and soft tissue inflammation, but previous radiation therapy was overlooked and the diagnosis was not made for several months after various tests, including biopsy. The two patients had different comorbidities and were managed by multidisciplinary collaboration between specialists. Surgery of the sacral ulcer was limited by the accessibility of non-irradiated tissues. In the absence of current guidelines, after radiopathological expertise, we used a "draining" procedure followed by antifibrotic pentoxifylline-tocopherol-clodronate treatment. CONCLUSION: Long after pelvic radiotherapy, symptomatic subcutaneous macrocalcification is suggestive of radiation-induced calcinosis. Prolonged antibiotic therapy followed by PENTOCLO treatment led to clinical improvement.

Loss of the disease-associated glycosyltransferase Galnt3 alters Muc10 glycosylation and the composition of the oral microbiome.

The importance of the microbiome in health and its disruption in disease is continuing to be elucidated. However, the multitude of host and environmental factors that influence the microbiome are still largely unknown. Here, we examined UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase 3 (Galnt3)-deficient mice, which serve as a model for the disease hyperphosphatemic familial tumoral calcinosis (HFTC). In HFTC, loss of GALNT3 activity in the bone is thought to lead to altered glycosylation of the phosphate-regulating hormone fibroblast growth factor 23 (FGF23), resulting in hyperphosphatemia and subdermal calcified tumors. However, GALNT3 is expressed in other tissues in addition to bone, suggesting that systemic loss could result in other pathologies. Using semiquantitative real-time PCR, we found that Galnt3 is the major O-glycosyltransferase expressed in the secretory cells of salivary glands. Additionally, 16S rRNA gene sequencing revealed that the loss of Galnt3 resulted in changes in the structure, composition, and stability of the oral microbiome. Moreover, we identified the major secreted salivary mucin, Muc10, as an in vivo substrate of Galnt3. Given that mucins and their O-glycans are known to interact with various microbes, our results suggest that loss of Galnt3 decreases glycosylation of Muc10, which alters the composition and stability of the oral microbiome. Considering that oral findings have been documented in HFTC patients, our study suggests that investigating GALNT3-mediated changes in the oral microbiome may be warranted.

Relationship of Streptococcus mutans with valvar cardiac tissue: A molecular and immunohistochemical study.

BACKGROUND: The present study aimed to investigate the presence or absence of Streptococcus mutans in oral cavity and valvular samples associating with the histomorphologic alterations of calcified aortic stenosis. METHODOLOGY: Dental plaque and cardiac valve samples were collected from 10 patients with calcified aortic stenosis for molecular analysis of S mutans by real-time polymerase chain reaction (PCR). Healthy valve tissue was also collected from five young cadavers and analyzed for S mutans. Moreover, fragments of all valvar specimens were submitted for histomorphological analysis and immunohistochemistry (anti-S mutans and anti-CD61). RESULTS: Streptococcus mutans was present in 100% of the oral cavity samples from the patients with calcified aortic stenosis in the molecular analysis. The analysis by real-time PCR showed that S mutans presented the same proportion in healthy valves and those with calcified aortic stenosis (80%; P = 1.000). Conversely, the immunoexpression of S mutans was 37.40 (IC95% = 1.49-937.00) times superior in samples of patients with cardiac disease (P = .007). The immunoexpression analysis showed that CD61 was present in seven (70%) calcified aortic stenosis samples, all of which were also immunopositive for S mutans. CONCLUSIONS: Streptococcus mutans was found in the oral cavity, healthy valve tissue, and calcified aortic stenosis samples. However, the microorganism was visualized by immunohistochemistry only in the calcified aortic stenosis samples, which may suggest viability and an increased bacterial density in this condition. The association of the presence of S mutans and positive CD61 immunoexpression suggests a probable relationship with calcified aortic stenosis.

Factors affecting bacterial culture positivity in specimens from bronchoscopy in patients with suspected lung cancer.

BACKGROUND: Bronchoscopy is important to diagnose lung cancer. However, some patients who undergo bronchoscopic procedures develop respiratory tract infections. Little is known about the proportion of pathogen-positive results in bacterial cultures from diagnostic bronchoscopy samples in patients with suspected lung cancer. This study aimed to determine the rate of positive bacterial cultures after diagnostic bronchoscopy in patients with suspected lung cancer and the relationship among culture results, clinical characteristics, and respiratory tract infections. METHODS: We retrospectively reviewed the medical records of all immunocompetent patients who underwent bronchoscopy and had culture and histological samples for the diagnosis of peripheral pulmonary lesions from September 2012 to August 2014 at the National Cancer Center in Tokyo. We analyzed data and classified radiological lesions into the following categories: calcifications, cavitations, low-density areas, margin irregularities, and satellite nodules. RESULTS: The study population consisted of 328 patients (median age, 69 years). We found that 65.9% of patients had malignant lesions and 4.2% of patients had positive cultures for pathogenic bacteria. The number of calcifications (p = 0.002, 95% CI: 2.17-41.10) was significantly higher in patients with positive bacterial isolates, according to the multivariate analysis, and bacterial culture positivity was not associated with the development of respiratory complications after bronchoscopy. Of the three patients with respiratory complications, all presented with cavitations. CONCLUSION: Because of the low prevalence of positive bacterial cultures in patients with suspected lung cancer, bacterial culture may be limited to specific patients, such as those with calcifications. Lesions with cavitation warrant close monitoring.

Morphological and micro-tomographic study on evolution of struvite in synthetic urine infected with bacteria and investigation of its pathological biomineralization.

Pathological biomineralization in the urinary system leads to urolithiasis. Formation of kidney stones involves a series of events during which they undergo morphological and mineralogical changes. We investigated the mineralization of biogenic struvite (in vitro) and examined the transformation of distinct interior and exterior structure of struvite. In vitro crystallization of struvite was performed in the presence of two bacteria that were originally isolated from the kidney stone patients. Morphological evaluation was carried out using SR-µCT as well as FESEM, XRD and FT-IR. Characteristic internal 3-D morphology and porosity of the stones were studied. For comparison, patient derived struvite stones were used. From the results obtained, we report that the presence of bacteria enhances the crystallization process of struvite in vitro. A series of time-resolved experiments revealed that struvite crystals experienced a significant morphologic evolution from pin pointed structure to X-shaped and tabular morphologies. These X-shaped and unusual tabular habits of struvite resembled biogenic morphologies of struvite. SR-µCT showed similarities between the patient derived and the in vitro derived struvite crystals. In conclusion, these experiments revealed that the bacteria play a major role in the specific morphogenesis of struvite and can able to control the nucleation, modulate crystalline phases, and shape of the growing crystal.

Prostate calcifications: A case series supporting the microbial biofilm theory.

Purpose: Prostate calcifications are a common finding during transrectal prostate ultrasound in both healthy subjects and patients, but their etiopathogenesis and clinical significance are not fully understood. We aimed to establish a new methodology for evaluating the role of microbial biofilms in the genesis of prostate calcifications. Materials and Methods: Ten consecutive patients who had undergone radical prostatectomy were enrolled in this study. All of the patients presented with prostate calcifications during transrectal ultrasound evaluation before surgery and underwent Meares-Stamey tests and clinical evaluation with the National Institutes of Health Chronic Prostatitis Symptom Index and the International Prostate Symptom Score. At the time of radical prostatectomy, the prostate specimen, after removal, was analyzed with ultrasonography under sterile conditions in the operating room. Core biopsy specimens were taken from the site of prostate calcification and subjected to ultrastructural and microbiological analysis. Results: The results of the Meares-Stamey test showed only 1 of 10 patients (10%) with positive cultures for Escherichia coli. Two of five patients (40%) had positive cultures from prostate biopsy specimens. Enterococcus faecalis, Enterococcus raffinosus, and Citrobacter freundii were isolated. Ultrastructural analysis of the prostate biopsy specimens showed prostate calcifications in 6 of 10 patients (60%), and a structured microbial biofilm in 1 patient who had positive cultures for E. faecalis and E. raffinosus. Conclusions: Although the findings are supported by a low number of patients, this study highlights the validity of the proposed methodology for investigating the role of bacterial biofilms in the genesis of prostate calcification.

Mitral Annular Calcification as a Possible Nidus for Endocarditis: A Descriptive Series with Bacteriological Differences Noted.

BACKGROUND: Mitral annular calcification (MAC) is a chronic inflammatory process with similarities to atherosclerosis. It is common in elderly patients and those with renal dysfunction. Although MAC is associated with cardiovascular morbidity, its relationship to infective endocarditis is unclear. The aim of this study was to test the hypothesis that MAC would be prevalent in patients with mitral valve vegetations and that vegetations would frequently occur on calcific nodules. A secondary aim was to look for possible bacteriological differences between vegetations attached to the calcified annulus versus leaflet vegetations. METHODS: We retrospectively reviewed all echocardiographic studies of patients with native mitral valve vegetations from January 2007 to August 2015 (N = 56). We searched for (1) presence of MAC, (2) location of MAC, and (3) vegetation location (on calcium deposits or distant). MAC was defined as focal echo brightness in a nodular or band-like pattern. The modified Duke criteria were used to confirm the diagnosis of infective endocarditis. Transthoracic, transesophageal, and three-dimensional echocardiograms (when available) at the time of infection were evaluated by a single reader. RESULTS: Twenty-eight subjects were infected with Staphylococcus aureus, 17 with a streptococcal species, and five with other organisms; blood cultures were sterile in 6. Thirty-four (61%) subjects had some degree of MAC, while 22 (39%) had none. Among those with MAC, the vegetation was located on the calcium deposits in 22 (65%), versus in 12 (35%) where it was not. Among all 56 subjects, when S. aureus was the infecting organism it was present on MAC in 16/28 (57%) versus 6/28 (21%; P = .01) for other bacterial species. By contrast, streptococcal infections more frequently involved the leaflets (16/17 [94%]) versus nonstreptococcal infections (18/39 [46%]; P = .0008). CONCLUSIONS: MAC may act as a nidus for infection especially with S. aureus. Differences in mechanism of attachment between S. aureus and streptococci may account for the observed difference in frequency of attachment of vegetations to MAC.

Prophylactic removal and microbiological evaluation of calcified plaques after pterygium surgery.

PURPOSE: The aim of this study was to investigate microbiological characteristics of prophylactically removed calcified plaques developed after pterygium excision, and to evaluate risk factors for the growth of microorganisms. METHODS: Only exposed calcified plaques developed at the same site of previous pterygium excision were prospectively removed in 15 eyes of 14 patients. Plaques were completely removed, divided into small pieces and evaluated for microbiological identification. Underlying scleral defects were reconstructed using a conjunctival autograft, amniotic membranes and scleral patch grafts according to the size and depth of the defects. Based on the results of microbiologic cultures, eyes were divided into two groups and risk factors for microbial growth were analyzed. RESULTS: At surgery, the mean age of the patients was 71.2 ± 5.8 years and 71.4 % were females. The mean time interval between pterygium excision and calcified plaque removal was 19.3 ± 13.8 years. Six of 15 (40 %) removed plaques showed bacterial growth, and Stenotrophomonas maltophilia was the most frequently isolated microorganism. The size of calcified plaques was the only risk factor for culture-positive results (p = 0.045). Underlying scleral defects were successfully repaired without any serious complication. CONCLUSIONS: Microorganisms can be isolated from calcified plaques developed at the site of previous pterygium excision, and the size of plaques is the only risk factor for culture-positive results. To remove potential source of infection, prophylactic removal of calcified plaques and scleral surface reconstruction should be considered, especially when the plaques are exposed and large.