[Changes of gut microflora in newly diagnosed IgA nephropathy patients and its correlation with clinical risk factors].

OBJECTIVE: To investigate the gut microbiota in newly diagnosed IgA nephropathy patients with chronic kidney disease (CKD) stages 1-2 and the association between the gut microbiota and the clinical risk factors of IgA nephropathy. METHODS: Fresh fecal samples were collected from nineteen newly diagnosed IgA nephropathy patients with CKD stages 1-2 and fifteen age- and sex-matched healthy controls. Fecal bacterial DNA was extracted and microbiota composition were characterized using 16S ribosomal RNA (16S rRNA) high-throughput sequencing for the V3-V4 region. The Illumina Miseq platform was used to analyze the results of 16S rRNA high-throughput sequencing of fecal flora. At the same time, the clinical risk factors of IgA nephropathy patients were collected to investigate the association between the gut microbiota and the clinical risk factors. RESULTS: (1) At the phylum level, the abundance of Bacteroidetes was significantly reduced (P=0.046), and the abundance of Actinobacteria was significantly increased (P=0.001). At the genus level, the abundance of Escherichia-Shigella, Bifidobacte-rium, Dorea and others were significantly increased (P < 0.05). The abundance of Lachnospira, Coprococcus_2 and Sutterella was significantly reduced (P < 0.05). (2) There was no significant difference in the abundance of gut microbiota between the newly diagnosed IgA nephropathy patients and the healthy control group (P>0.05), but there were differences in the structure of the gut microbiota between the two groups. The results of LEfSe analysis showed that there were 16 differential bacteria in the newly diagnosed IgA nephropathy patients and healthy controls. Among them, the abundance of the newly diagnosed IgA nephropathy patients was increased in Enterobacteriales, Actinobacteria, Escherichia-Shigella, etc. The healthy control group was increased in Bacteroidetes and Lachnospira. (3) The result of redundancy analysis (RDA) showed that Bifidobacterium was positively correlated with serum IgA levels, 24-hour urinary protein levels and the presence of hypertension. Lachnoclostridium was positively correlated with the presence of hypertension. Escherichia-Shigella was positively correlated with urine red blood cells account. Bifidobacterium was positively correlated with the proliferation of capillaries. Faecalibacterium was positively correlated with cell/fibrocytic crescents. Ruminococcus_2 was positively correlated with mesangial cell proliferation, glomerular segmental sclerosis and renal tubular atrophy/interstitial fibrosis. CONCLUSION: The gut microbiota in the newly diagnosed IgA nephropathy patients with CKD stages 1-2 is different from that of the healthy controls. Most importantly, some gut bacteria are related to the clinical risk factors of IgA nephropathy. Further research is needed to understand the potential role of these bacteria in IgA nephropathy.

[Comparison of clinical features and high-resolution esophageal motility characteristics between esophagogastric junction outflow obstruction and type â ¡ achalasia patients].

OBJECTIVES: To compare the clinical features and high-resolution esophageal motility-impedance characteristics among esophagogastric junction outflow obstruction (Eoo) patients, type Ⅱ achalasia (Ach) patients and healthy controls (Con), in order to explore the values of esophageal high-resolution manometry (HRM) in diagnosis and treatment of Eoo patients. METHODS: Patients with dysphagia were enrolled from December 2011 to December 2014 at the outpatient department of Peking University Third Hospital, so were age-matched healthy volunteers. All the patients with organic obstruction were excluded. All the participants were tested with high-resolution esophageal motility-impedance measurement, the patients were diagnosed as Eoo or Ach according to the Chicago classification criteria. Clinical features and esophageal motility characteristics of Eoo, Ach and Con were analyzed. RESULTS: A total of 23 Eoo, 24 Ach and 20 Con were enrolled, whose gender ratios, average ages and body mass indexes were of no significant differences(all P>0.05). (1) The Eoo group had higher percentage of food reflux[21.7% (5/23) vs 0(0/24), P=0.005]and belching[17.4% (4/23) vs 0 (0/24), P=0.013], but lower percentage of dysphagia[47.8% (11/23) vs 79.2% (19/24), P=0.025]and vomiting[0(0/23) vs 12.5%(3/24), P=0.040]compared with the Ach group, with no significant differences in other symptoms(all P>0.05). Besides, the Eoo group had lower Eckardt scores than the Ach group[1(1, 2) vs 3 (2, 5), P<0.001]. (2) The lower esophageal sphincter (LES) basal pressure-minimum in the Eoo was higher than the Con[(26.73±2.77) vs (17.16±1.76) mmHg, P<0.05]. The mean LES basal pressure; and the LES integrated relaxation pressure (IRP), IRP-maximum, and LES relaxation percentage were significantly different among Eoo, Ach and Con[(19.80±1.25) vs (35.95±2.36), (8.43±0.72) mmHg, both P<0.05; (23.22±2.02) vs (48.37±3.71), (12.32±1.29) mmHg, bothP<0.05; 38.61%±3.10% vs 12.42%±5.66%, 64.00%±3.85%, both P<0.05]. (3) There were significant differences in velocity, amplitude, and duration of esophageal peristaltic wave and intrabolus pressure (all P<0.05) among Eoo, Ach and Con; and failed contraction percentage, panesophageal pressurization percentage, premature contraction percentage, and rapid contraction percentage of Eoo were lower than Ach (all P<0.05) while complete contraction percentage of Eoo was high compared with Ach (P<0.001), but no significant differences between Eoo and Con. (4) The Eoo had significantly less incomplete bolus clearance[0.00% (0.00%, 20.00%) vs 100.00% (90.00%, 100.00%), P<0.001]and shorter bolus transit time[(5.44±0.29) s vs (24.13±1.69) s, P<0.001]than Ach, but there were no significant differences between Eoo and Con in these two indexes[0.00% (0.00%, 20.00%) vs 0.00% (0.00%, 9.75%); (5.44±0.29) s vs (5.30±0.19) s; both P>0.05]. CONCLUSIONS: The clinical manifestations and esophageal HRM characteristics of Eoo appear to be between Ach and Con, which suggests that Eoo may be an early-stage of Ach. Further study of the pathophysiological characteristics of Eoo patients may provide more evidence to elucidate the pathogenesis of achalasia.

Establishment of cell clones with different metastatic potential from the metastatic hepatocellular carcinoma cell line MHCC97.

AIM: To establish clone cells with different metastatic potential for the study of metastasis-related mechanisms. METHODS: Cloning procedure was performed on parental hepatocellular carcinoma (HCC) cell line MHCC97, and biological characteristics of the target clones selected by in vivo screening were studied. RESULTS: Two clones with high (MHCC97-H) and low (MHCC97-L) metastatic potential were isolated from the parent cell line. Compared with MHCC97-L, MHCC97-H had smaller cell size (average cell diameter 43 microm vs 50 microm) and faster in vitro and in vivo growth rate (tumor cell doubling time was 34.2h vs 60.0h). The main ranges of chromosomes were 55-58 in MHCC97-H and 57-62 in MHCC97-L. Boyden chamber in vitro invasion assay demonstrated that the number of penetrating cells through the artificial basement membrane was (37.5 +/- 11.0) cells/field for MHCC97-H vs (17.7 +/- 6.3)/field for MHCC97-L. The proportions of cells in G0-G1 phase, S phase, and G2-M phase for MHCC97-H/MHCC97-L were 0.56/0.65, 0.28/0.25 and 0.16/0.10, respectively, as measured by flow cytometry. The serum AFP levels in nude mice 5wk after orthotopic implantation of tumor tissue were (246 +/- 66) microg.L(-1) for MHCC97-H and (91 +/- 66) microg.L(-1) for MHCC97-L. The pulmonary metastatic rate was 100% (10/10) vs 40% (4/10). CONCLUSION: Two clones of the same genetic background but with different biological behaviors were established, which could be valuable models for investigation on HCC metastasis.

[Expression of alpha-smooth muscle actin in scar fibroblasts in vitro].

OBJECTIVE: To explore the expression of alpha-smooth muscle actin (alpha-SMA) induced by transforming growth factor beta 1 (TGF-beta 1). METHODS: Five samples of hypertrophic scars and three samples of normal mature scars were collected as the experimental and control groups respectively. The fibroblasts were isolated from scars, and cultured in 2-dimension or 3-dimension culture system. The immunohistochemical staining method of LSAB were used to investigate the expression of alpha-SMA in fibroblasts in the different concentration of TGF-beta 1. RESULTS: The expression of alpha-SMA in 3-dimension culture system were markedly lower than those in 2-dimension culture system with respect to the fibroblasts in the experimental group. The expression of alpha-SMA in fibroblasts were different in response to various TGF-beta 1 concentration, it was more effective at the concentration of 5 ng/ml. The expression of alpha-SMA in the fibroblasts from hypertrophic scars seemed to be more sensitive to TGF-beta 1 compared to that of the normal mature scars. CONCLUSION: There are concentration-dependent in the expression of alpha-SMA induced by TGF-beta 1 in scar fibroblasts in vitro. The biological characteristics of the fibroblasts from hypertrophic scars and normal mature scars and their sensitivity to the inducement of TGF-beta 1 were different. The inducement of TGF-beta 1 may be depressed by extracellular matrix components and that may decrease the expression of alpha-SMA.

[Changes in muscle of a muscle flap after severance of its innervating nerve. A preliminary study].

The purpose of this study is to observe changes in muscle after severance of its main innervation, especially the changes in its volume and weight. In ten rabbits, it was found that the reduction of the volume of gastrocnemius muscle flap was 47.8%, and that of gastrocnemius and soleus muscles was 50.5% one month after the severance of the tibial nerve, while their weights were decreased 47.6%, 51.3% and 46.9% respectively. After three months the respective reduction in volume was 40.3%, 32.3% and 38.8%, while the weight was reduced to 44.0%, 34.0 and 40.3%, respectively. Histological examination revealed capillary dilatation and congestion, decrease myofibrils and increase of adipose tissue. Evidence of diminution of muscular metabolism and energy consumption was found by histochemical study, including ATPase, phosphorylase, and succinic dehydrogenase staining. The result of electromyogram examination confirms the presence of muscle denervation.

[Growth of transplanted auricular cartilage in young rabbits].

Three months after isotopic and heterotopic autotransplantation of 100 pieces of auricular cartilage grafts (10 x 10 mm each) in 25 seven-week-old rabbits, growth was observed in 93% of the grafts. The length increased 24.0% to 26.0%, the width 12.0% to 17.0%, the area 40.0% to 47.4%. A synchronous growth of the transplant and the ear was observed with the growth of the rabbits. Histologic study showed that the growth of transplanted auricular cartilage is achieved jointly by two mechanisms, namely interstitial growth and appositional growth. The results of this experiment offer a positive and encouraging outlook for autotransplantation of cartilage in young patients.

[Experimental research on absorption after ilium autografting].

Even though efforts were made by closely adhering to the established rules of bone grafting, late absorption of bone grafts occurs frequently. Sometime the effect of operation is jeopardized. The cause of bone absorption is complicated, it is necessary to carry out further research. In this study 80 autogenous grafts were taken from ilium of 40 rabbits. A half of the grafts were transplanted on to cranium, the rests were implanted into subcutaneous tissue. One month later, bone grafts were absorbed 20.0% (on surface of cranium bone) and 21.0% (in subcutaneous sites). Three months after operation absorptions increased to 67.6% and 69.1% respectively. Six months later, absorption reached to 76.0% and 73.6%. There is no statistical difference between two sites where the grafts are transplanted (P greater than 0.05). Absorption of bone grafts was also found in 3 dogs. Its ilium autografts were absorbed 52.8% (over cranium) and 45.2% (in subcutaneous tissue) 3 months after operation, the former increased to 55.3% and the latter 50.1% 6 months later. Microscopic examination revealed absorption and melting of bone grafts, necrosis and cysts formation were also found in some grafts. We believe that these data may help us to recognize the absorption further after free bone autografting.