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Objective Nanobacteria are one of the factors for urinary calculi and its exact pathogenic mechanism is not yet clear.The aim of this study was to investigate the role of the CaSR -Claudin-14 regulatory channel in the formation of calculi . Methods Sixty Wistar male rats were equally randomized into a normal control group and nanobacterial group , the former injected via the tail vein with 1.2 mL of 0.9％ sodium chloride solution while the latter with 1.2 mL of nanobacterial suspension , both for once.Three of the rats in each group were sacrificed every week in the first 10 weeks after injection.Histopathological examination was performed every week to evaluate the stone formation in the kidneys of the rats , and the expressions of the CaSR and Claudin -14 proteins were determined by immunohistochemistry. Results From the 1st to the 10th week after injection, crystal particles were observed in the rat kidneys of the nanobacterial group, but not in the normal controls (52.4％ vs 0％, P<0.01).The expressions of CaSR and Claudin -14 showed no statistically significant differences between the nanobacterial and control groups in the first 3 weeks (P>0.05) but both gradually in-creased in the former group from the 4th to the 10th week as compared with the latter, mainly in membrane of the renal tubular epithelial cells. Conclusion The increased activity of the CaSR -Claudin-14 regulatory channel may play an important role in the formation of nanobacterial renal stone .
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Objective To observe the early effect of Fructus mume extract on KIM-1 and OPN levels in rats with kidney stone formation induced by nanobacteria and to investigate the therapeutic significance of F.mume extract on early kidney stone formation.Methods Nanobacteria were separated and cultured from human upper urinary calculi.The study group appropriately included 54 healthy male SD rats.The renal calculus model was constructed by tail vein injection of nanobacteria.A kidney stone model was created with an F.mume extract intervention,and rats were killed at different stages of the intervention.Real-time polymerase chain reaction was used to detect the KIM-1 mRNA expression in rat renal tissue,and the KIM-1 concentration in urine was detected by using enzyme-linked immunosorbent assay.We detected kidney tissue stone crystals and OPN expression by using hematoxylin-eosin staining and immunohistochemistry.Results Renal tubules of the experimental model were significantly expanded,that is,the formation of renal tubular stones.The early KIM-1 and OPN expression levels were increased.The above-mentioned changes positively correlated with the injection time of the nanobacteria,and F.mume extract antagonized the changes.Conclusion F.mume extract may be useful for the repair of renal injury to reduce kidney stone formation,which may be related to the gene regulation of KIM-1 and OPN.
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Objective To investigate the clinical efficacy of Dahuang Xiaozhi Suppository combined with tetracycline in treating type Ⅲ prostatitis infected with nano-bacteria. Methods Totally 120 patients of type Ⅲprostatitis infected with nano-bacteria were randomly divided into treatment group and control group, with 60 cases in each group. Both groups were disabled anti-infective drugs and other preparations for diet and life intervention. Both groups received tetracycline, once a tablet, twice a day, orally. The treatment group received Dahuang Xiaozhi Suppository, once a capsule, once a day, placing in the anus 3-4 cm. 10 d was a treatment course, for 3 courses. The clinical efficacy, major symptoms improving time, NIH-CPSI, leukocyte count and ecithin corpuscles in expressed prostatic secretion (EPS), urinary flow rate, and cytokine content of TNF-α, IL-6, IL-8, were evaluated. Results The total effective rate was 100.0% (60/60) in treatment group and 83.3% (50/60) in the control group, and treatment group was higher than the control group (P<0.05). Pelvic pain, urinary symptom, and scrotum wet improvement time of treatment group were lower than the control group (P<0.05). Compared with before the treatment, the NIH-CPSI pain scores, urinary symptom scores, life quality score, leukocyte count and TNF-α, IL-6, IL-8 content of EPS in both groups were significantly lower after treatment (P<0.05), and the cases with lecithin corpuscles +++ - ++++ in both groups significantly increased after treatment (P<0.05). The urinary peak flow rate and mean flow rate in both groups were higher after treatment (P<0.05). There was statistical significance in the scores of NIH-CPSI pain, urinary symptom and life quality, leukocyte count and TNF-α, IL-6, IL-8 content of EPS, and the cases with lecithin corpuscles ++++ between the two groups (P<0.05). Conclusion Dahuang Xiaozhi Suppository can improve the efficacy of treating type Ⅲ prostatitis infected with nano-bacteria. Combining Dahuang Xiaozhi Suppository with tetracycline can reduce prostate inflammation, with obvious efficacy.
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Objective To investigate the infection status of nanobacteria on patients with upper urinary calculi and healthy sub-jects ,and analyze the role of nanobacteria in the formation of upper urinary calculi .Methods The serum ,urine and calculus of 42 patients with upper urinary calculi were investigated by polymerase chain reaction (PCR) and by bacteria cultivation with 10% γ-FBS and PMBI 1640 .The resulting PCR products were sequenced for further comparison with the reported sequence in gene bank . The serum and urine from 30 healthy adults were used as controls .Results After 4 to 6 weeks′culture ,the white or yellow precipi-tate was visible at the bottom of the tube .The positive rate of PCR was 90 .4% in calculous patients urine and 6 .7% in healthy a-dults urine ,as the positive was 92 .8% and 6 .7% in serum .which there is significant difference (P<0 .01) .The positive rate of the nanabactria in urinary calculi was 95 .2% .The coincidence rate was 98 .72% between the PCR products and the reported sequence in gene bank .Conclusion Nanobacteria are widely existed in the serum ,urine and calculus of the patients with upper urinary calcu-li ,this indicate that the nanobacteria might be have the most important influence on the formation process of urinary calculi .
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Objective To reproduce an SD rat model of prostatic calculus by using nanobacteria (NB), and explore the role of NB in contributing to prostatitis and prostatic calculus. Methods Twenty adult male SD rats were randomized to the control group and 20 to the model group. Rat prostate infection models were reproduced by infusing 0. 2 ml (Concentration, 1 Mai unit) NB suspension transurethrally. 0.2 ml physiological saline was infused transurethrally in the rat control group. The rats were sacrificed 4 and 8 weeks later and prostatic pathology were viewed by hematoxylin and eosin (HE) staining. Lithogenesis was observed by scanning electron microscope (SEM) or Transmission electron microscopy (TEM). Re-isolation, culture and identification of nanobacteria were also done in rat prostatic tissues. Results Chronic inflammatory changes of prostates were shown in the model group at both 4 weeks and 8 weeks after infusing NB suspension. Prostatic calculi were detected by SEM and TEM at 8 weeks in the prostates of the rat model group (7/10). Neither chronic inflammatory changes nor prostatic calculus was found in the control group. NB was positive in the model group, but negative in the control group. Conclusions NB infection could cause chronic prostatitis and prostatic calculus in rats.
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Objective To identify the nanobaeteria in prostate fluid of patients with CPPS.Methods Expressed prostatic secretion(EPS)and urine specimens were collected by Meares-Stamey way from CPPS patients(n=100)and normal controls(n=100).The specimens were cultured and nanobacteria was identified by indirect immunofluoreseenee staining with rnonoelonal antibody.The morphological features were observed by using transmission electron microscopy(TEM). Results The positive rate of nanobaeteria in the EPS cdture of CPPS patients and controls were 43% and 5% respectively,with significant statistical difference(X2=39.58,P＜0.01).By TEM,the sizes of NB ranged from 100 to 500 nm and appeared eoccoid-ccccobacillary shape. Conclusion Nanobaeteria infection may exist in EPS of CPPS patients.
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Objective To investigate the infection status of nanobacteria on patients of chronic hepatopathy and hepatocellular carcinoma,and evaluate the clinical value of PCR.Methods In sera of 68 cases of chronic hepatitis B(CHB),56 chronic severe hepatitis B(CSHB),66 cirrhosis of liver(CL)and 23 hepatocellular carcinoma(HCC),nanobacteria were detected by immunohistochemistry stain(IHC),Transmission Electron Microscopy(TEM)and polymerase chain reaction(PCR),compared with 40 healthy people.Results The positive rates of PCR were 27.69%,50.00%,61.29%,52.38% and 5.00% in patients with CHB,CSHB,CL,HCC and normal control respectively(P
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Objective:To study the damage of nanobacteria on HK-2 cells,the possible principles,the effect of crystals(COM) adhering to HK-2 cells after the damage.Methods:Four groups were chosen for the study:control group,NB group,nHAP group and COM group.Morphological changes of the HK-2 cells were observed after HE stain and with TEM after 12 hours and 24 hours.Meanwhile,the levels of H2O2,LDH,MDA and ATPases were surveyed after 6 hours,12 hours and 24 hours,respectively.And 6,12,and 24 hours later,COM crystals were mixed into the culture fluids of each group.Then phalloidin-FITC was used to finish fluorescent staining of the cells.At last,the adhering effects of each group with the laser scanning confocal fluorescence microscope were observed and contrasted.Results:After HE stain and with TEM:in NB and nHAP group,the shape of the cells changed,brush borders were arranged in disorder,vacuoles formed in the kytoplasms,the mitochondria became swelled up,the karyotheca dissolved and the nucleolus disappeared in some cells.After 24 hours,in NB group,the number of the cells in which the karyotheca dissolved was more than that in nHAP group.After 12 and 24 hours,the level of H2O2 in NB group was higher than that in control group and nHAP group;After 6 and 24 hours,the level of MDA in NB group was higher than that in control group and nHAP group;At each time point,there was no significant difference in the level of LDH between control group,nHAP group and NB group;After 12 hours,the activities of Na+/K+ ATPases in NB group and nHAP group were lower than those in control group.And after 24 hours,the activity of Na+/K+ ATPases in NB group was lower than that in control group;After 12 and 24 hours,the activities of Ca2+ /Mg2+ ATPases in NB group was lower than those in control group.After 12 hours,the activity of Ca2+/Mg2+ ATPases in nHAP group was lower than that in control group.The observation with the laser scanning confocal fluorescence microscope:after 12 hours,showed that the number of the crystals adhering to the cells in NB group and COM group increased,and in COM group,some crystals had entered the cells;after 24 hours,the adhering effects of the crystals in NB and COM group were similar to those after 12 hours,but the number of adhered crystals was more than that after 12 hours;At each time point,there was no significant change in control and nHAP groups.Conclusion:Nanobacteria has a damage effect on HK-2 cells,the damage increases with the acting time expanding.The damage is more severe than that of nHAP.In the damage process of nanobacteria,the lipid peroxidation may play an important role.After the damage of nanobacteria,the adhering effect of the COM crystals to the cells increases observably,and the number of crystals adhering to the cells becomes more and more with the acting time expanding.Although nHAP also has a damage effect on HK-2 cells,it does not effect the adhering process.
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Objective:To detect,culture,and characterize the nanobacteria(NB) from sera of patients with kidney calculi in our department.Methods:Blood samples of 24 patients with kidney calculi and of 3 healthy volunteers in our department were collected for NB culture in this study.We used immunohistochemistry,von kossa staining,scanning electron microscopy(SEM),transmission electron microscopy(TEM) to investigate the appearance and components of cultural NB.Results:Twenty-two blood samples out of 24(91.67%) showed growth of NB,while no NB were detected in volunteers' blood samples.The infection rate of stone group was obviously higher than that of healthy volunteers.After a 4-week culture period,the light microscope revealed coccoid-shaped NB with a diameter of 100-500 nm,which could be identified by immunohistochemistry and von kossa staining.SEM and TEM(negative staining) revealed NB with a hollow interior coated in needle-like apatite crystals.Such nanopaticles could bud-off new ones and therefore appeared like living organisms.Conclusion:NB can be identified from sera of most patients involved in kidney calculi.It may have intimate relation to the formation of kidney calculi because the infection rate of NB blood samples of stone patients was significantly higher than that of healthy volunteers.Immunohistochemistry,von kossa staining,SEM and TEM are special methods for identifying NB from different aspects.The appearance and character are important points to distinguish NB from other nano-sized particles.
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Objective To investigate the chronic inflammation causing effect of nanobacteria(NB) on the prostates of SD rats,and to provide experiment evidence for etiology of type Ⅲ prostatitis.Methods Forty adult,male rats were divided into model group(16 rats),treatment group(8 rats) and control group(16 rats) at random.The rats in model group and treatment group were reproduced by infusing NB suspension transurethrally,and the animals of control group received an infusion of normal saline.After 4 weeks,WBC,lecithine and pathological changes of prostates were observed in model group(8 rats) and control group(8 rats).Tetracycline(100 mg?kg-1?d-1) was administered to treatment group,and distilled water were given to other groups respectively for 4 weeks.Then all rats were sacrificed,and the same detections were done as before.NB from prostates of animal were re-isolated,cultured and identified.Results After 4 weeks,chronic inflammation were observed in model group.WBC became significantly higher and lecithine of prostates were lower in model group than in control group(P0.05).The positive cases of NB were 16 in model group,and none in control group.Conclusion Nanobacteria causes chronic inflammation in SD rat prostates.NB maybe the etiological agent of type Ⅲ prostatitis.Anti-NB treatment by tetracycline is effective for this inflammation.
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Objective To explore the distribution of the nanobacteria in the prostatic calculus.Methods Calculus specimens were collected from 40 patients of prostatic calculus during operation.After the stones were crushed and rinsed with saline,the liquid was diluted,filtrated and centrifuged,then cultured in strict aseptic condition.The obtained cells were observed using transmission electron microscopy(TEM),detected with alizarin red staining and Gram staining,and identified by indirect immunofluorescence staining with monoclonal antibody 8D10 against nanobacteria.Results ①After 3 to 4 weeks' culture,a white precipitate adhering to the tube bottom was found.②The positive rate of nanobacteria were 65.0% in 40 calculus patients.③Alizarin red staining and Gram staining indicated nanobacteria were distributed in cluster.④Nanobacteria appeared coccoid or coccobacillary-like in shape and ranged from 100 to 500 nm in size.Conclusion Nanobacteria infection exists in calculus patients.
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Nanobacteria (NB) is a kind of new bacteria with a diameter of 8 0~500 nm. It has specific mineralizing ability. As a active nidus it can attac h, invade and damage the renal epithelium of collecting ducts and papilla, and t hen form apatite which being the center to induce formation of kidney stones. I n the paper, the research progress on nanobateria contained in kidney stones and its role in kidney stones formation were summarized. The simulation in vitro a nd animal models of kidney stones formation induced by nanobateria were discusse d.
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ObjectiveTo explore the distribution of nanobacteria in prostatic calculus and investigate its role in the formation of prostatic calculus. MethodsThe stones of 40 patients with prostatic calculus was used to isolate and culture the possible bacteria. The genomes of obtained bacteria were extracted, and the 16S rRNA was amplified by PCR followed by sequencing. ResultsThe obtained specific fragment had a 98% resemblance with 16S rRNA of nanobacteria: Score=2 480 bits (1 290), Expect=0.0; Identities=1 387/1 409 (98%), Gaps=4/1 409 (0%); Strand=Plus/Plus. ConclusionNanobacteria is proved existing in the stones of prostatic calculus patients by PCR and sequencing.