Precise analysis of the effect of basal core promoter/precore mutations on the main phenotype of chronic hepatitis B in mouse models.

High replication and mutation rates of hepatitis B virus (HBV) often lead to reduced or suppressed hepatitis B e antigen expression. The most common mutations are genomic variations in the basal core promoter (BCP) and pre-core (PC) regions. However, the effect of BCP/PC mutations on HBV phenotype in vivo remains unclear. We compared and analyzed BCP/PC mutations and BCP/PC reverse mutations in mouse models. In addition to terminating the expression of HBeAg, BCP/PC mutations also resulted in a significant decrease in HBsAg, HBV DNA, and cccDNA in the early stage, and an obvious increase in serum alanine aminotransferase throughout the transfection period. In both groups, serum HBV DNA was positively correlated with intracellular HBV DNA and cccDNA. Further, we found that interleukin-4 (IL-4) and L-10 levels were significantly lower in the BCP/PC(M) group than in the BCP/PC(R) group at 4 weeks post-injection. However, IL-1ß was significantly lower in the BCP/PC(M) group than in the BCP/PC(R) group at 26 weeks post-injection. In summary, we precisely analyzed the effect of BCP/PC mutations on the phenotype in vivo, which is important to evaluating disease progression and treatment responses of variable chronic hepatitis B patients.

Spectroscopic Study on the Interaction between Naphthalimide-Polyamine Conjugates and Bovine Serum Albumin (BSA).

The effect of a naphthalimide pharmacophore coupled with diverse substituents on the interaction between naphthalimide-polyamine conjugates 1-4 and bovine serum albumin (BSA) was studied by UV absorption, fluorescence and circular dichroism (CD) spectroscopy under physiological conditions (pH = 7.4). The observed spectral quenching of BSA by the compounds indicated that they could bind to BSA. Furthermore, caloric fluorescent tests revealed that the quenching mechanisms of compounds 1-3 were basically static type, but that of compound 4 was closer to a classical type. The Ksv values at room temperature for compound-BSA complexes-1-BSA, 2-BSA, 3-BSA and 4-BSA were 1.438 × 104, 3.190 × 104, 5.700 × 104 and 4.745 × 105, respectively, compared with the value of MINS, 2.863 × 104 at Ex = 280 nm. The obtained quenching constant, binding constant and thermodynamic parameter suggested that the binding between compounds 1-4 with BSA protein, significantly affected by the substituted groups on the naphthalene backbone, was formed by hydrogen bonds, and other principle forces mainly consisting of charged and hydrophobic interactions. Based on results from the analysis of synchronous three-dimensional fluorescence and CD spectra, we can conclude that the interaction between compounds 1-4 and BSA protein has little impact on the BSA conformation. Calculated results obtained from in silico molecular simulation showed that compound 1 did not prefer either enzymatic drug sites I or II over the other. However, DSII in BSA was more beneficial than DSI for the binding between compounds 2-4 and BSA protein. The binding between compounds 1-3 and BSA was hydrophobic in nature, compared with the electrostatic interaction between compound 4 and BSA.

Study on the synthesis, biological activity and spectroscopy of naphthalimide-diamine conjugates.

Eleven novel naphthalimide-diamine conjugates were synthesized and their structures were confirmed by elemental analysis, 1H-NMR, 13C-NMR and MS. Their in vitro antitumor activities were assessed using MTT assays on two cancerous cell lines K562, HCT116, and one normal hepatoma cell line QSG 7701. Compound 7f exhibited potent antitumor activity on HCT116 cells and favorable cell selectivity toward QSG 7701 compared with the positive control, amonafide. Moreover, 7f could block HeG2 cells in the G2/M phase and induce HeG2 cells apoptosis. The interaction of compound 7f with herring sperm DNA was studied by UV/vis absorption and fluorescence spectroscopy under physiological conditions (pH = 7.4). The observed spectral quenching of compound 7f by DNA and the displacement of EB from DNA-EB complex by compound 7f indicated that compound 7f could intercalate into DNA base pairs, which was also corroborated by the effect of KI on compound-DNA interaction. Further caloric fluorescent tests revealed that the quenching mechanism was a static type. Meanwhile, the binding constants, thermodynamic parameters and the effect of NaCl on compound-DNA interaction showed that the type of interaction force was mainly hydrogen bonds and the binding process was driven by hydrogen and van der Waals bonding.

Sarcopenia adversely impacts clinical outcomes in patients undergoing pancreaticoduodenectomy: A systematic review and meta-analysis.

BACKGROUND: Sarcopenia is a syndrome marked by a gradual and widespread reduction in skeletal muscle mass and strength, as well as a decline in functional ability, which is associated with malnutrition, hormonal changes, chronic inflammation, disturbance of intestinal flora, and exercise quality. Pancreatoduodenectomy is a commonly employed clinical intervention for conditions such as pancreatic head cancer, ampulla of Vater cancer, and cholangiocarcinoma, among others, with a notably high rate of postoperative complications. Sarcopenia is frequent in patients undergoing pancreatoduodenectomy. However, data regarding the effects of sarcopenia in patients undergoing pancreaticoduodenectomy (PD) are both limited and inconsistent. AIM: To assess the influence of sarcopenia on outcomes in patients undergoing PD. METHODS: The PubMed, Cochrane Library, Web of Science, and Embase databases were screened for studies published from the time of database inception to June 2023 that described the effects of sarcopenia on the outcomes and complications of PD. Two researchers independently assessed the quality of the data extracted from the studies that met the inclusion criteria. Meta-analysis using RevMan 5.3.5 and Stata 14.0 software was conducted. Forest and funnel plots were used, respectively, to demonstrate the outcomes of the sarcopenia group vs the non-sarcopenia group after PD and to evaluate potential publication bias. RESULTS: Sixteen studies encompassing 2381 patients were included in the meta-analysis. The patients in the sarcopenia group (n = 833) had higher overall postoperative complication rates [odds ratio (OR) = 3.42, 95% confidence interval (CI): 1.95-5.99, P < 0.0001], higher Clavien-Dindo class ≥ III major complication rates (OR = 1.41, 95%CI: 1.04-1.90, P = 0.03), higher bacteremia rates (OR = 4.46, 95%CI: 1.42-13.98, P = 0.01), higher pneumonia rates (OR = 2.10, 95%CI: 1.34-3.27, P = 0.001), higher pancreatic fistula rates (OR = 1.42, 95%CI: 1.12-1.79, P = 0.003), longer hospital stays (OR = 2.86, 95%CI: 0.44-5.28, P = 0.02), higher mortality rates (OR = 3.17, 95%CI: 1.55-6.50, P = 0.002), and worse overall survival (hazard ratio = 2.81, 95%CI: 1.45-5.45, P = 0.002) than those in the non-sarcopenia group (n = 1548). However, no significant inter-group differences were observed regarding wound infections, urinary tract infections, biliary fistulas, or postoperative digestive bleeding. CONCLUSION: Sarcopenia is a common comorbidity in patients undergoing PD. Patients with preoperative sarcopenia have increased rates of complications and mortality, in addition to a poorer overall survival rate and longer hospital stays after PD.

Exosome-Transmitted miR-224-5p Promotes Colorectal Cancer Cell Proliferation via Targeting ULK2 in p53-Dependent Manner.

Objective: To investigate the role and molecular mechanism of exosomal miR-224-5p in colorectal cancer (CRC). Methods: The miR-224-5p expression in CRC patient tissues and cell-derived exosomes was measured by laser capture microdissection and qRT-PCR, respectively. Dual-luciferase reporter gene assay was used to determine the target gene of miR-224-5p. The protein expressions of p53 and unc-51 like kinase 2 (ULK2) in CRC cells were detected by western blot. Flow cytometry was used to detect cell cycle and apoptosis. Cell proliferation was measured by CCK8 and EdU assay. Results: The miR-224-5p expression was upregulated in CRC tissues and increased progressively with the rise of CRC stage. CRC cells secreted extracellular miR-224-5p mainly in an exosome-dependent manner, and then miR-224-5p could be transferred to surrounding tumor cells to regulate cell proliferation in the form of autocrine or paracrine. Moreover, ULK2 was characterized as a direct target of miR-224-5p and was downregulated in CRC tissues. Interestingly, ULK2 inhibited CRC cell proliferation in a p53-dependent manner. Furthermore, exosome-derived miR-224-5p partially reversed the proliferation regulation of ULK2 on CRC cells. Conclusion: Our findings demonstrate that exosome-transmitted miR-224-5p promotes p53-dependent cell proliferation by targeting ULK2 in CRC, which may offer promising targets for CRC prevention and therapy.

254C>G: a TRPC6 promoter variation associated with enhanced transcription and steroid-resistant nephrotic syndrome in Chinese children.

BACKGROUND: Mutations in canonical transient receptor potential channel 6 (TRPC6) have been identified as responsible for the development of focal segmental glomerulosclerosis, a proteinuric disease with steroid resistance and poor prognosis. This study explores the prevalence of TRPC6 variants in Chinese children with idiopathic nephrotic syndrome (INS), the genotype/phenotype correlation of TRPC6 variants, the therapeutic response, and the underlying molecular mechanism. METHODS: Fifty-one children with sporadic INS were enrolled: 23 steroid-sensitive cases and 28 steroid-resistant cases Polymerase chain reaction was used to amplify 13 exons and the promoter sequences of TRPC6 before sequencing. The expression of TRPC6 in renal tissues was illustrated by immunohistochemistry staining. The transcriptional activity of variants in TRPC6 promoter was measured by the luciferase assay. RESULTS: Three variants (-254C>G, rs3824934; +43C/T, rs3802829; and 240 G>A, rs17096918) were identified. The allele frequency of the -254C>G single-nucleotide polymorphism (SNP) in the steroid-resistant nephrotic syndrome (SRNS) patients (40.5%) was higher than that in the steroid-sensitive nephrotic syndrome subjects (27.1%; P = 0.046). The -254C>G SNP enhanced transcription from TRPC6 promoter in vitro and was associated with increased TRPC6 expression in renal tissues of SRNS patients. CONCLUSION: -254C>G, a SNP underlying enhanced TRPC6 transcription and expression, may be correlated with the development of steroid resistance in Chinese children with INS.

[Expression of neonatal Fc receptor on human nephritis and rat nephritis models].

OBJECTIVE: To study the expression of neonatal Fc receptor in podocytes in human nephritis and immune-induced rat nephritis models: anti-Thy1.1 nephritis and Heymann nephritis. METHODS: Thirty-nine cases of renal biopsies were enrolled from September 2009 to February 2010, including 8 cases of minimal change disease, 4 cases of focal segmental glomerulosclerosis, 9 cases of membranous nephropathy, 12 cases of IgA nephropathy and 6 cases of lupus nephritis. Five normal kidney tissue samples adjacent to renal clear-cell carcinoma were served as normal controls. Laser capture microdissection and real-time RT-PCR were used to assess the expression level of FcRn mRNA in glomeruli of various glomerulonephritides, and immunohistochemistry (IHC) of FcRn by SuperVision method was performed. In addition, rat models of mesangial proliferative nephritis (anti-Thy1.1 nephritis) and passive membranous nephropathy (Heymann nephritis) were established and FcRn was examined in renal tissues by IHC. RESULTS: The FcRn mRNA level in lupus nephritis was statistically higher than that of normal controls (P < 0.05). FcRn protein expression by IHC was seen in lupus nephritis (6/6), membranous nephropathy (6/9) and IgA nephropathy (7/12), significantly higher than that of normal controls (0/5), P < 0.05. Minimal change disease and focal segmental glomerular sclerosis showed minimal or none expression of FcRn (1/8, 0/4 respectively) and not statistically difference from that of normal controls. Furthermore, FcRn expression in podocytes was detected in rat anti-Thy1.1 (3/5) and Heymann nephritis models (2/7) but was not detected in normal controls. CONCLUSIONS: Expression of FcRn in podocytes was up-regulated in immune-induced human nephritis and rat nephritis models of anti-Thy1.1 nephritis and Heymann nephritis. FcRn may play a role in the development of immune-induced glomerulonephritis.

[Historical Changes and Responses to Human Activities of Polycyclic Aromatic Hydrocarbons in Lake Sediments from Northern China During the Past 100 Years].

Polycyclic aromatic hydrocarbons (PAHs) are of concern globally because of their carcinogenic, teratogenic, mutagenic, and bio-accumulative effects. Northern China is one of the regions in China with a high density of lakes; however, the lake aquatic environment is becoming seriously deteriorated, especially from PAH pollution due to the intensification of human activities during the past 100 years. Therefore, the spatial distribution and historical changes in PAHs in lake sediments from northern China were analyzed to indicate their response to anthropogenic emissions and pollution reduction actions. The ω(PAHs) in lake sediments ranged from 18.2 to 1205.0 ng·g-1, and low molecular weight PAHs were the dominant compounds. PAH concentrations increased from the 1950s to a peak level in the 2000s, which was induced mainly by increased energy consumption and rapid economy development, with PAH levels decreasing subsequently in the last 10 years due to craft improvement of wastewater treatment plants and the promotion of new energy policies. Spatially, PAHs pollution in Northeast and North China was more serious than that in Northwest China due to the higher level of economic development and energy consumption. Source apportionment results revealed that historical PAH emissions transferred from biomass combustion to a mixture of coal and petroleum combustion. In addition, the results of ecological risk assessment showed that the synthetic sediment quality index (SeQI) of northern China ranged from 36 to 75, and North and Northeast China posed higher ecological risk than that in Northwest China, with phenanthrene (Phe), acenaphthylene (Ace), acenaphthylene (Acy), and dibenzo(a,h)anthracene (DahA) as the main risk contributors.

Expression of response gene to complement-32 in renal tissue of children with immunoglobulin A nephropathy.

OBJECTIVE: This study aimed to investigate the expression and significance of response gene to complement-32 (RGC-32) in renal tissue of children with immunoglobulin A nephropathy (IgAN). MATERIAL AND METHODS: Forty-five patients diagnosed as having IgAN by renal biopsy were enrolled. The expression of RGC-32, α-smooth muscle actin (α-SMA) and transforming growth factor-ß(1) (TGF-ß(1)) was observed by immunohistostaining. The relationshis between the expression of RGC-32, α-SMA, TGF-ß1, degree of renal pathological lesions in IgAN and clinical index were assessed by Spearman correlation. RESULTS: Immunohistostaining analysis showed that RGC-32 protein was present in epithelial cells of renal tubules in normal and IgAN renal tissues. With more severe renal pathological lesions, the expression of RGC-32 in IgAN was increased. The expression of RGC-32 was positively correlated with that of α-SMA, TGF-ß(1) and the degree of renal pathological lesions in children with IgAN (p < 0.05), but had no relationship with serum creatinine, urinary N-acetyl-ß-d-glucosaminidase/creatinine, urinary microalbuminuria/creatinine, urinary microimmunoglobulin/creatinine or urinary α(1)-microglobulin/creatinine ratio (p > 0.05). CONCLUSION: Expression of RGC-32 can reflect the degree of renal pathological lesions in IgAN. RGC-32 may participate in the renal tubulointerstitial lesions in children with IgAN, especially in epithelial -mesenchymal transition induced by TGF-ß(1).

[Effects of Chinese herbal medicine Huaiqihuang Granule on nephrin and podocin expressions in renal tissues of rats with adriamycin-induced nephrosis].

OBJECT: To investigate the effects of Huaiqihuang Granule, a compound Chinese herbal medicine, on expressions of nephrin and podocin of slit diaphragm of glomerular podocytes in rats with adriamycin-induced nephrosis and to explore the mechanism in reducing the proteinuria. METHODS: Twenty SD rats were randomly divided into five groups: control group, model group, glucocorticoid group, Huaiqihuang Granule group and Huaiqihuang Granule plus glucocorticoid group. The 24-hour urine was collected 7, 14, 21 and 28 days after adriamycin injection respectively to measure 24-hour urinary protein, and all rats were sacrificed after 28-day treatment. Pathological changes in renal tissues were observed under a light microscope and an electron microscope. Expressions of nephrin and podocin mRNAs in renal cortex were determined by real-time polymerase chain reaction, and protein levels of nephrin and podocin were detected by Western blotting. RESULTS: (1) In the model group and the treatment groups, the level of urinary protein increased significantly from the 14th day. (2) Under the light microscope, inflammatory cells and slight fibroplasia were found in renal interstitium of the model group, but there were less inflammatory cells in renal interstitium in the intervention groups than in the model group. Under the electron microscope, 29 days after adriamycin injection, extensive fusion of foot processes was observed. (3) The expressions of nephrin and podocin were higher in treatment groups than in the model group. (4) Proteinuria level was negatively correlated with the expressions of nephrin mRNA and nephrin and podocin proteins. CONCLUSION: The above results indicate that Huaiqihuang Granule can maintain the integrity of the slid diaphragram in podocyte, alleviate the lesion of glomerular filtration membrane, and decrease the proteinuria by up-regulating the expressions of nephrin and podocin. Huaiqihuang Granule plus glucocorticoid maybe has better effects than glucocorticoid alone.

[Screening of Priority Pollutants and Risk Assessment for Surface Water from Shengjin Lake].

Shengjin Lake, which serves as an important National Nature Reserve, is suffering from chemical pollution due to rapid industrial and agricultural development in the circumjacent basin. Therefore, 168 anthropogenic toxic chemicals were determined to examine their spatial distribution and identify priority pollutants using a ranking system based on occurrence(O), persistence(P), bioaccumulation(B), ecological risk(E), and human health risk(H). Ecosystem and human health risks were also assessed. The spatial distribution of pollutants indicated that higher concentrations occur in the upper lake area compared to the middle and lower lake areas because of Jiang Dam. According to the derived priority pollutant list, phthalate esters(PAEs), organochlorine pesticides(OCPs), and heavy metals(HMs) are high-priority pollutants; polychlorinated biphenyls(PCBs), polycyclic aromatic hydrocarbons(PAHs), and volatile organic compounds(VOCs) are medium-priority pollutants; and antibiotics(ANTs) are low-priority pollutants. The ecology risk quotient(RQ) of the high-priority pollutants ranged from 4.3 to 15.9, indicating severe ecology risk to the aquatic organism, and higher risks were found in the upper lake areas. Additionally, the human health risk assessment revealed negligible carcinogenic risks associated with high-priority pollutants. The comprehensive ranking system established in this study can be applied to other lake basins by altering the measured concentrations to screen for priority pollutants, offering a scientific foundation for identifying priority control pollutants for watershed management.

[Variations of Stable Oxygen and Deuterium Isotopes in River and Lake Waters During Flooding Season Along the Middle and Lower Reaches of the Yangtze River Regions].

Stable isotope tracers have been widely applied to water sources and evolution, transforming relations, and pollution sources of various water bodies. This study analyzed the spatial variations of δ2H and δ18O in river and lake waters during flooding season, and revealed the factors underlying their variations along the middle and lower reaches of the Yangtze River based on a field sampling campaign in July 2018. Our results showed that δ2H and δ18O in the Yangtze River water were enriched from the Three Gorges reservoir region to the lower reaches of the Yangtze River, which was closely linked to isotopic variations in precipitation. There was no significant difference in δ2H and δ18O values in the mainstream river waters between the Three Gorges Reservoir Region and Yichang-Chenglingji. However, d-excess values in river water displayed a small variation range. In contrast, δ2H and δ18O values in the lake group from Dongting to Jianghan and Huayang to Poyang Lake were lower than in the lake group from Taihu to the Yangtze Delta. Negative d-excess values were observed in lake water from Taihu to the Yangtze Delta, suggesting the combined influence of enriched isotopic compositions in precipitation and strong evaporative enrichment. Of the lakes, the highest isotopic values were found in Dianshan Lake and Datong Lake, whereas the lowest isotopic values were recorded in Dongting Lake and Poyang Lake because of their direct connection with the Yangtze River. The water regimes of Dongting Lake and Poyang Lake were influenced by the Yangtze River, especially when a high water level of the Yangtze River occurred, and thus altered the isotopic compositions of Dongting Lake and Poyang Lake water. Hence, these findings will provide scientific data revealing the precipitation-river-lake interactions and investigating the rational utilization and management of water resources in the middle and lower reaches of the Yangtze River regions.

The Effects of Neurokinin-1 Receptor Antagonist in an Experimental Autoimmune Cystitis Model Resembling Bladder Pain Syndrome/Interstitial Cystitis.

To identify the effects of the neurokinin-1 receptor (NK1R) antagonist aprepitant in treating pelvic pain, micturition symptoms, and bladder inflammation in mice with experimental autoimmune cystitis (EAC) similar to bladder pain syndrome/interstitial cystitis (BPS/IC). Female C57BL/6 mice were divided into the following three groups: normal control, EAC, and EAC plus aprepitant. EAC was induced in mice by duplicate immunization with bladder homogenate. In the EAC model group, EAC mice were given PBS by gavage once a day during the fourth week. In the EAC plus aprepitant group, aprepitant was administered instead of PBS in the same way. After 4 weeks, pelvic pain threshold and urination habits of mice were analyzed, as well as the bladder weight to body weight ratio, and histologic assessment of the expression of IL-1ß, TNF-α, intercellular adhesion molecule 1 (ICAM-1), and NK1R in bladder tissue. EAC mice mimicked the phenotype and pathophysiologic lesions of BPS/IC well. Compared to PBS-treated EAC mice, the mice treated with aprepitant exhibited higher pain threshold values, less number of total urine spots or small urine spots, lower bladder weight to body weight ratio, and reduced bladder inflammation with less mast cell infiltration and decreased expressions of IL-1ß, TNF-α, and ICAM-1 in bladder tissue. There was no difference in NK1R expression in bladders treated with or without aprepitant. The NK1R antagonist aprepitant relieved pelvic pain, urinary symptoms, and bladder inflammation in EAC mice. This indicated that NK1R may be a novel therapeutic target in BPS/IC treatment.

[Effect of tissue factor pathway inhibitor on apoptosis of rat mesangial cells and Fas and bcl-2 expression].

OBJECTIVE: To study the biological impact and mechanism of recombinant tissue factor pathway inhibitor (rTFPI) on apoptosis of rat kidney mesangial cells (MsC). METHODS: TFPI expression in human glomerular minor lesion (GML), mesangial proliferative glomerulonephritis (MPGN) and cultured rat MsC was detected using immunohistochemistry and immunofluorescence, respectively. Rat MsC were incubated with rTFPI and its variant peptides. Morphological changes of apoptosis were investigated by Hoechst 33258 and the apoptotic rate was assessed by flow cytometry. DNA fragmentation and effect of rTFPI on expression of caspase-3, Fas and bcl-2 were studied using gel electrophoresis and Western blot respectively. RESULTS: The expression of TFPI in MPGN was higher than that in GML. TFPI was expressed in cultured rat mesangial cells. Apoptosis of MsC was induced by rTFPI, especially by its C-termianl, in a dose- and time-dependent manner. Apoptosis ratios of MsC treated with rTFPI were 2.1, 3.0 and 4.9 times more than control, respectively. Expression of gene caspase-3 and Fas was up-regulated in a dose-dependent manner wherease bcl-2 expression did not show any changes. CONCLUSION: rTFPI induces apoptosis in cultured rat mesangial cells by its C-terminal possibly via Fas/FasL pathway.

The effect of TGF-ß1 and Smad7 gene transfer on the phenotypic changes of rat alveolar epithelial cells.

The aim of this study was to investigate whether transforming growth factor-ß1 (TGF-ß1) could induce alveolar epithelial-mesenchymal transition (EMT) in vitro, and whether Smad7 gene transfer could block this transition. We also aimed to elucidate the possible mechanisms of these processes. The Smad7 gene was transfected to the rat type II alveolar epithelial cell line (RLE-6TN). Expression of the EMT-associated markers was assayed by Western Blot and Real-time PCR. Morphological alterations were examined via phase-contrast microscope and fluorescence microscope, while ultrastructural changes were examined via electron microscope. TGF-ß1 treatment induced a fibrotic phenotype of RLE-6TN with increased expression of fibronectin (FN), α-smooth muscle actin (α-SMA) and vimentin, and decreased expression of E-cadherin (E-cad) and cytokeratin19 (CK19). After transfecting the RLE-6TN with the Smad7 gene, the expression of the mesenchymal markers was downregulated while that of the epithelial markers was upregulated. TGF-ß1 treatment for 48 h resulted in the separation of RLE-6TN from one another and a change into elongated, myofibroblast-like cells. After the RLE-6TN had been transfected with the Smad7 gene, TGF-ß1 treatment had no effect on the morphology of the RLE-6TN. TGF-ß1 treatment for 48 h resulted in an abundant expression of α-SMA in the RLE-6TN. If the RLE-6TN were transfected with the Smad7 gene, TGF-ß1 treatment for 48 h could only induce a low level of α-SMA expression. Furthermore, TGF-ß1 treatment for 12 h resulted in the degeneration and swelling of the osmiophilic multilamellar bodies, which were the markers of type II alveolar epithelial cells. TGF-ß1 can induce alveolar epithelial-mesenchymal transition in vitro, which is dependent on the Smads signaling pathway to a certain extent. Overexpression of the Smad7 gene can partially block this process.

[Effect of Smad7 on transforming growth factor-beta1-induced alveolar epithelial to mesenchymal transition].

OBJECTIVE: To investigate whether transforming growth factor beta1 (TGF-beta1) can induce in vitro alveolar epithelial-mesenchymal transition (EMT), and whether Smad7 gene transfer can block this transition and the possible signaling mechanism. METHODS: Rat alveolar type II epithelial cells of the line RLE-6TN were cultured. TGF beta1 (3 ng/mL) was added into the culture fluid. Lipofectamine 2000 was used to transfect Smad7 gene to the RLE-6TN cells. The expression of the markers of the epithelial cells, including E-cadherin and cytokeratin-19 (CK19), and markers of mesenchymal cells, including fibronectin (FN), vimentin, and alpha-smooth muscle actin (alpha-SMA) were assayed using Western blotting and real-time PCR. The morphological alterations were examined by phase-contrast microscope while the ultrastructure changes were examined by electron microscope. RESULTS: Smad7 was successfully transfected to the RLE-6TN cells. Before transfection TGF-beta1 treatment could lead to the expression upregulation of the mesenchymal markers and downregulation of the epithelial markers at the levels of both mRNA and protein, and after transfection, the mesenchymal makers were downregulated while the epithelial markers were upregulated. Before the transfection, TGF-beta1 treatment could lead to the expression upregulation of phosphorylated Smad2/3 which did not obvious change following transfection. TGF beta1 treatment could induce the EMT process of the RLE-6TN cells. Smad7 gene transfected into the RLE-6TN cells could block the process of EMT. Exposed of the RLE-6TN cells to TGF beta1 resulted in degeneration, tumefaction, and gradual disappearance of the osmiophilic multilamellar bodies, markers of type II alveolar epithelial cells. CONCLUSION: Under TGF beta1 treatment, RLE-6TN undergoes a conversion process into myofibroblasts in vitro with the conversion mechanism related to Smad signaling pathway, and transfection of Smad7 gene can partly reverse this process.

[Clinicopathologic analysis of 34 patients with microscopic polyangitis].

OBJECTIVE: To study the clinicopathologic features of microscopic polyangitis (MPA), and to compare the differences in anti-neutrophil cytoplasmic autoantibody (ANCA)-positive and ANCA-negative patients, as well as in ANCA-positive cases with or without glomerular immunoglobulin deposition. METHODS: Thirty-four biopsy-proven cases of MPA were retrieved from the archival files of the Department during the past 7 years. The clinicopathologic characteristics between ANCA-positive and negative patients, as well as between ANCA-positive cases with and without glomerular immunoglobulin deposition, were compared. RESULTS: Amongst the 34 MPA patients studied, about one-fifth to one-half were accompanied by various extrarenal symptoms. Serum ANCA was positive in 26 patients (76.5%). A slight to moderate increase in urinary protein was demonstrated in 31 patients, while 3 patients had nephrotic syndrome. Elevated serum creatinine was detected in 32 cases. Renal biopsy revealed crescentic glomerulonephritis in 24 cases, focal segmental glomerulonephritis in 8 cases, vascular fibrinoid necrosis with inflammation in 7 cases, intimal thickening of arterioles in 24 cases, interstitial inflammatory cells, including neutrophil infiltration (21 cases), in 29 cases. Crescentic formation was more common in the ANCA-positive group than in the ANCA-negative group (P < 0.05). Amongst the 26 ANCA-positive cases, 10 had glomerular immunoglobulin deposits (including 1 case with IgA nephropathy). In general, these cases had a greater degree of proteinuria than those without glomerular immunoglobulin deposits (P < 0.05). CONCLUSIONS: The diagnosis of MPA relies on histologic examination of renal biopsy and clinicopathologic correlation. Serum ANCA seems important for glomerular crescent formation. Glomerular immunoglobulin deposition may also play a significant role in the exacerbation of proteinuria.

Remediation of PCB-contaminated soil using a combination of mechanochemical method and thermal desorption.

The combination of mechanochemical method and thermal desorption for remediating polychlorinated biphenyls (PCBs) in contaminated soil was tested in this study. The effects of grinding time and heating time on PCB removal efficiency were investigated. The contaminated soil, mixed with CaO powder at a weight ratio of 1:1, was first ground using a planetary ball mill. After 4 h of grinding, the total PCB concentration and its toxic equivalence quantity (TEQ) decreased by 74.6 and 75.8%, respectively. Then, after being heated at 500 °C for 60 min, the residual PCBs in mechanochemical + thermal treated soil decreased to 247 ng/g, resulting in a removal efficiency of 99.95%. The removal effect can be promoted by longer grinding time and heating time; however, increased energy consumption was inevitable. The combination of grinding time and heating time should be optimized in a practical remediation process.

Establishment of a Novel Autoimmune Experimental Model of Bladder Pain Syndrome/Interstitial Cystitis in C57BL/6 Mice.

The aim of this study is to identify whether vaccinating twice with bladder homogenate can establish a new model of experimental autoimmune cystitis (EAC) in C57BL/6 strain mice. C57BL/6 mice were vaccinated with bladder homogenate in complete Freund's adjuvant (CFA) and boost immunized with bladder homogenate in incomplete Freund's adjuvant (IFA) after 2 weeks were used as the EAC model. Mice immunized with phosphate-buffered saline (PBS) in CFA or IFA were used as the control. Micturition habits and suprapubic-pelvic pain threshold were measured 4 weeks after primary immunization. Bladder to body weight ratios and expression of inflammatory cytokines and neurokinin 1 receptor (NK1R) were then examined. Histologic and immunohistochemical examination of the bladder was carried out, and IL-1ß, IFN-γ, and TNF-α production by the kidneys, liver, and lungs was also tested. Double-immunized mice were extensively sensitive to pressure applied on the pelvic area (P < 0.001). Compared to single-immunized mice or controls, double-immunized mice showed more micturition frequency, lower urine output per micturition, higher bladder to body weight ratio, and significant elevation in the expression of inflammatory cytokines, including IL-1ß, IL-4, IL-6, IL-10, IFN-γ, and TNF-α (all P < 0.05). NK1R gene expression was significantly increased in double-immunized mice compared to the other three groups (P < 0.001). A nonspecific immune response occurred in the liver but was much weaker than bladder inflammation. Our dual immunization EAC model in C57BL/6 mice can effectively mimic the symptoms and pathophysiologic characteristics of BPS/IC and thus can be widely used to investigate the pathogenesis and therapeutic strategies of BPS/IC.

A new model mimicking persistent HBV e antigen-negative infection using covalently closed circular DNA in immunocompetent mice.

Despite the availability of an effective vaccine, hepatitis B virus (HBV) infection remains a major health problem. HBV e antigen (HBeAg)-negative strains have become prevalent. Previously, no animal model mimicked the clinical course of HBeAg-negative HBV infection. To establish an HBeAg-negative HBV infection model, the 3.2-kb full-length genome of HBeAg-negative HBV was cloned from a clinical sample and then circularized to form covalently closed circular (cccDNA). The resulting cccDNA was introduced into the liver of C57BL/6J mice through hydrodynamic injection. Persistence of the HBeAg-negative infection was monitored at predetermined time points using HBV-specific markers including HBV surface antigen (HBsAg), HBeAg, and HBV core antigen (HBcAg) as well as DNA copies. Throughout the study, pAAV-HBV1.2 was used as a control. In mice injected with HBeAg-negative cccDNA, the HBV infection rate was 100% at the initial stage. HBsAg levels increased up to 1 week, at which point levels peaked and dropped quickly thereafter. In 60% of injected mice, HBsAg and HBcAg persisted for more than 10 weeks. High numbers of HBV DNA copies were detected in the serum and liver. Moreover, cccDNA persisted in the liver tissue of HBeAg-negative mice. In contrast to the pAAV-HBV 1.2 injected mice, no HBeAg was found in mice injected with HBeAg-negative HBV throughout the study period. These results demonstrate the first successful establishment of a model of HBeAg-negative HBV-persistent infection in immunocompetent mice. Compared to pAAV-HBV1.2-injected mice, the infection persistence and levels of serum virological and biochemical markers were approximately equal in the model mice. This model will be useful for mechanistic studies on HBeAg-negative HBV infection and will facilitate the evaluation of new antiviral drugs.