Background At present,a new drug,platelet-derived growth factor-BB (PDGF-BB),as a drug knife for the treatment of glaucoma is under study to expect it directly working on the trabecular meshwork without disrupting the normal physiological structure of anterior chamber angle,clearing the trabecular meshwork aqueous outflow channel so as to achieve the purpose of lowering intraocular pressure.Objective This study aimed to investigate the effect of PDGF-BB on matrix metalloproteinase-2 (MMP-2) expression in cultured bovine trabecular meshwork cells.Methods Trabcular tissue was obtained from fresh bovine eyes,and trabcular meshwork cells were cultured and passaged using explant method.Cultured cells were identified by morphological evaluation and neuronspecific enolase (NSE) staining.The third generation of cells were inoculated to 6-well plate,and different concentrations (0,5.0,12.5,25.0 μg/L) of PDGF-BB was added into the medium for 2 hours.Expression levels (A value) of MMP-2 mRNA (MMP-2 mRNA/β-actin) and protein in the cells were assayed by reverse transcription polymerase chain reaction (RT-PCR) and immunochemistry,respectively.Results Trabcular meshwork cells appeared 5-9 days after cultured.The third generation of cells presented with many process and showed the green influence in cytoplasm.MMP-2 mRNA/β-actin value (A) was 0.127 ± 0.026,0.147 ± 0.045,0.178 ± 0.053 and 0.222±0.062 in the O,5.0,12.5,25.0 μg/L PDGF-BB group,respectively,showing a significant difference among them (F =56.71,P＜0.05),and the MMP-2 mRNA/β3-actin value in the 5.0,12.5,25.0 μg/L PDGF-BB group was elevated in comparison with that of the 0 μg/L PDGF-BB group (all P＜0.05).The expression value (A value) of PDGF-BB protein in the cells was 446.12±13.81,1444.65±54.64,2086.18±73.18,3488.65±25.98 in the 0,5.0,12.5,25.0 μg/L PDGF-BB group,respectively,with a significant difference among the four groups (F=213.12,P＜0.01),and the expression value (A value) of PDGF-BB protein was gradually increased with the ascend of concentration of PDGF-BB(all P＜0.05).Conclusions PDGF-BB can promote the expression of MMP-2 in bovine trabcular meshwork cells in vitro in concentration-dependent manner.

OBJECTIVETo study the inhibitory effect of Xiaotan Sanjie Recipe (XSR) on the microsatellite instability of orthotopic transplantation tumor in MKN-45 human gastric cancer nude mice.

METHODSThe 3rd passage subcutaneous transplantation tumor was taken as the origin of the model by using MKN-45 human gastric cancer cell lines. MKN-45 human gastric cancer nude mouse model was established using OB glue adhesive method. Then 30 nude mice were divided into the model group, the XSR group, and the chemotherapy group. Mice in the XSR group were intragastrically given XSR at the daily dose of 0.4 mL. Mice in the chemotherapy group were intragastrically given Fluorouracil at the daily dose of 0.4 mL. No intervention was given to mice in the model group. After 6 weeks of medication, the tumor weight was measured, and the tumor inhibition rate calculated. The size, the peak height, and the peak area of 5 microsatellite instability sites were detected.

RESULTSThe tumor inhibition rate was 40. 84% in the XSR group. The tumor weight was significantly lower in the XSR group than in the model group (P < 0.01), showing no statistical difference when compared with the chemotherapy group (P >0.05). The incidence of high microsatellite instability (MSI-H) in the model group was 70%, and the incidence of low microsatellite instability (MSI-L) was 30%. Microsatellite stable site tended be stable after 6 weeks of XSR treatment.

CONCLUSIONXSR showed inhibition on microsatellite instable orthotopic transplantation tumor in MKN-45 human gastric cancer nude mice.

Animals ; Antineoplastic Agents ; pharmacology ; Cell Line, Tumor ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Mice ; Mice, Nude ; Microsatellite Instability ; drug effects ; Neoplasm Transplantation ; Stomach Neoplasms

Purpose: We investigated the association of effector memory (EM) CD8 + T cell and CD4 + T cell immunity with metabolic syndrome (MS). Methods: Surface and intracellular staining of peripheral blood mononuclear cells was performed. Anti-interleukin-7 receptor-alpha (IL-7Rα) and CX3CR1 antibodies were used to stain the subsets of EM CD8 + T cells, while anti-interferon-gamma (IFN-γ), interleukin-17 (IL-17), and forkhead box P3 (FOXP3) antibodies were used for CD4 + T cell subsets. Results: Of the 47 obese children, 11 were female. Children with MS had significantly higher levels of serum insulin (34.8±13.8 vs. 16.4±6.3 μU/mL, p<0.001) and homeostasis model assessment of insulin resistance (8.9±4.1 vs. 3.9±1.5, p<0.001) than children without MS.Children with MS revealed significantly higher frequencies of IL-7Rα low CD8+ T cells (60.1 ±19.1% vs. 48.4±11.5%, p=0.047) and IL-7Rα low CX3CR1 + CD8 + T cells (53.8±20.1% vs. 41.5 ±11.9%, p=0.036) than children without MS. As the serum triglyceride levels increased, the frequency of IL-7Rα low CX3CR1 + and IL-7Rα high CX3CR1 – CD8 + T cells increased and decreased, respectively (r=0.335, p=0.014 and r=−0.350, p=0.010, respectively), in 47 children. However, no CD4 + T cell subset parameters were significantly different between children with and without MS. Conclusion In obese children with MS, the changes in immunity due to changes in EM CD8 + T cells might be related to the morbidity of obesity.

Purpose: We investigated the association of effector memory (EM) CD8 + T cell and CD4 + T cell immunity with metabolic syndrome (MS). Methods: Surface and intracellular staining of peripheral blood mononuclear cells was performed. Anti-interleukin-7 receptor-alpha (IL-7Rα) and CX3CR1 antibodies were used to stain the subsets of EM CD8 + T cells, while anti-interferon-gamma (IFN-γ), interleukin-17 (IL-17), and forkhead box P3 (FOXP3) antibodies were used for CD4 + T cell subsets. Results: Of the 47 obese children, 11 were female. Children with MS had significantly higher levels of serum insulin (34.8±13.8 vs. 16.4±6.3 μU/mL, p<0.001) and homeostasis model assessment of insulin resistance (8.9±4.1 vs. 3.9±1.5, p<0.001) than children without MS.Children with MS revealed significantly higher frequencies of IL-7Rα low CD8+ T cells (60.1 ±19.1% vs. 48.4±11.5%, p=0.047) and IL-7Rα low CX3CR1 + CD8 + T cells (53.8±20.1% vs. 41.5 ±11.9%, p=0.036) than children without MS. As the serum triglyceride levels increased, the frequency of IL-7Rα low CX3CR1 + and IL-7Rα high CX3CR1 – CD8 + T cells increased and decreased, respectively (r=0.335, p=0.014 and r=−0.350, p=0.010, respectively), in 47 children. However, no CD4 + T cell subset parameters were significantly different between children with and without MS. Conclusion In obese children with MS, the changes in immunity due to changes in EM CD8 + T cells might be related to the morbidity of obesity.

OBJECTIVETo explore a method of extracting tumor interstitial fluid (TIF) which is similar to muddy phlegm in Chinese medicine (CM), interleukin-8 (IL-8) in concentration was taken as the representative of the content of TIF, analyzed in the extracted TIF and the original tumor tissue, and examined to see whether TIF has an interfering effect on tumor recurrence.

METHODSTumor tissue was ground, centrifuged, and filtered for intercellular substances. Tumor-bearing Kunming S180 mice were raised for 21 days and then the tumors were removed to observe the influence of intervention with TIF, normal saline (NS) and a blank control on tumor recurrence.

RESULTSThe content of IL-8 in the filtered and unfiltered tumor tissue was not significantly different (P>0.05). Postoperative tumor recurrence in TIF intervention group was significantly higher than that in the NS intervention and control groups (60%, 12/20 vs. 20%, 4/20. vs. 15%, 3/20, χ(2) =11.058, P<0.01). Tumor cells grew vigorously and infiltrated to muscular tissue in TIF intervention group. Large numbers of tumor cells were seen necrotic in the NS intervention group, and small numbers of tumor cells were seen necrotic in the blank control group.

CONCLUSIONSTIF can be effectively extracted by the means described. It does not contain tumor cells, but its contents such as IL-8 may stimulate tumor cell growth and promote postoperative tumor recurrence, which provided preliminary experimental basis for hypothesis of "tumor-phlegm microenvironment".

Animals ; Enzyme-Linked Immunosorbent Assay ; Extracellular Fluid ; Interleukin-8 ; analysis ; Mice ; Neoplasms, Experimental ; pathology ; Postoperative Period ; Recurrence

OBJECTIVETo extract tumor interstitial fluid (TIF) from MKN-45 gastric cancer which is similar to "muddy phlegm" in Chinese medicine and observe influences of MKN-45 tumor interstitial fluid (MKN-45 TIF) intervention on metastasis of gastric cancer and on the expressions of vascular endothelial growth factor (VEGF), kinase insert domain containing receptor (KDR), epithelial-cadherin (E-cad), cyclooxygenase-2 (COX-2), intercellular adhesion molecule-1 (ICAM-1) and telomerase genes and proteins in primary tumor tissue.

METHODSAn MKN-45 tumor-bearing model was established in 50 nude mice. The modeled animals were equally randomized to 5 groups: the simple tumor-bearing group (model group), the normal saline (NS) via tail vein injection (i.v.) group (NS i.v. group), MKN-45 TIF i.v. group (TIF i.v. group), NS intraperitoneal injection (i.p.) group (NS i.p. group), and MKN-45 TIF i.p. group (TIF i.p. group). The TIF and NS intervention groups received injection (i.p. or i.v.) of MKN-45 TIF or NS twice a week, 0.2 mL at a time. After 8 weeks, the primary tumors were removed, weighed and HE stained to observe tumor metastasis. The primary tumor tissues were analyzed by immunohistochemistry and real-time quantitative PCR to detect expressions of VEGF, KDR, E-cad, COX-2, ICAM-1, and telomerase genes and proteins in different groups.

RESULTSThere were significant differences in tumor weight between TIF intervention groups and the model and NS intervention groups. Tumor metastasis was observed in all 5 groups, but the tumor metastasis rate in TIF intervention groups was significantly higher than those in the model and NS intervention groups. The gene and protein expressions of gastric cancer-related factors VEGF, KDR, COX-2, ICAM-1 and telomerase were unregulated while the gene and protein expressions of E-cad were downregulated in TIF intervention groups.

CONCLUSIONSTIF promotes tumor growth, invasion and metastasis of gastric cancer. These findings provide preliminary experimental clues for verifying the hypothesis of "tumor-phlegm microenvironment".

Animals ; Cadherins ; genetics ; metabolism ; Cell Line, Tumor ; Cyclooxygenase 2 ; genetics ; metabolism ; Extracellular Fluid ; metabolism ; Gene Expression Regulation, Neoplastic ; Humans ; Intercellular Adhesion Molecule-1 ; genetics ; metabolism ; Male ; Mice ; Mice, Nude ; Neoplasm Transplantation ; Stomach Neoplasms ; metabolism ; secondary ; Telomerase ; genetics ; metabolism ; Tumor Microenvironment ; physiology ; Vascular Endothelial Growth Factor A ; genetics ; metabolism ; Vascular Endothelial Growth Factor Receptor-2 ; genetics ; metabolism

OBJECTIVETo elucidate the roles of JAK/STATs signal pathway on anti-proliferative effects induced by IFN-alpha in MHCC97.

METHODSAn IRF9 expression vector was transfected into MHCC97 with Dosper. The expression of IRF9, cycle regulating proteins and the forming of ISGF3 complex were detected using Western blot and EMSA, respectively. Cell proliferation and distribution were monitored using MTT and flow cytometry.

RESULTSHigh expression of IRF9 restored the anti-proliferative response of MHCC97 on IFN-alpha treatment and delayed the cell transition from S phase to G2 phase induced by IFN-alpha.

CONCLUSIONThe integrity and functions of JAK/STATs signal pathway played an important role in mediating the anti-proliferative effects of IFN-alpha in MHCC97.

Carcinoma, Hepatocellular ; genetics ; metabolism ; pathology ; Cell Line, Tumor ; Cell Proliferation ; Humans ; Interferon-Stimulated Gene Factor 3, gamma Subunit ; genetics ; Interferon-alpha ; metabolism ; pharmacology ; Janus Kinases ; genetics ; physiology ; Liver Neoplasms ; genetics ; metabolism ; pathology ; STAT Transcription Factors ; genetics ; physiology ; Signal Transduction ; Transfection

Objectives: Previous studies have suggested that the lactic acid bacterium, Weissella cibaria CMU has beneficial effects on halitosis, but its precise effects have not been evaluated in human subjects. We evaluated the efficacy and safety of W. cibaria CMU for reducing halitosis in adults (20-70 years old) whose exhibited volatile sulfur compound (VSC) concentrations exceeded 0.015 ng/mL and who scored ≥2 points in a halitosis sensory evaluation test. Methods: A total of 60 participants were assigned to an experimental group (treated with W. cibaria CMU) and a control group (placebo). In total, 58 out of 60 participants (experimental group, 29; control group, 29) were ultimately included in gas chromatography (OralChroma) analyses of VSC concentrations and halitosis sensory evaluation tests. Results: We found that the VSC concentration decreased by 0.030±0.062 ng/ml in the experimental group after 8 weeks (P=0.0138) and increased by 0.005±0.124 ng/ml in the control group (P=0.8198). However, the difference between groups was not statistically significant (P>0.05). In a sensory evaluation test, a significantly lower score was obtained for the experimental group than for the control group. Conclusions Overall, VSC concentrations and sensory evaluation scores were lower in the experimental group than in the control group, but only the latter was statistically significant. Thus, we conclude that W. cibaria CMU is involved in the reduction of halitosis.

Background and Objectives: Brain organoids have the potential to improve our understanding of brain development and neurological disease. Despite the importance of brain organoids, the effect of vascularization on brain organoids is largely unknown. The objective of this study is to develop vascularized organoids by assembling vascular spheroids with cerebral organoids. Methods: and Results: In this study, vascularized spheroids were generated from non-adherent microwell culture system of human umbilical vein endothelial cells, human dermal fibroblasts and human umbilical cord blood derived mesenchymal stem cells. These vascular spheroids were used for fusion with iPSCs induced cerebral organoids. Immunostaining studies of vascularized organoids demonstrated well organized vascular structures and reduced apoptosis. We showed that the vascularization in cerebral organoids up-regulated the Wnt/β-catenin signaling. Conclusions We developed vascularized cerebral organoids through assembly of brain organoids with vascular spheroids. This method could not only provide a model to study human cortical development but also represent an opportunity to explore neurological disease.

The high percentage of Vermamoeba was found in tap water in Korea. This study investigated whether Vermamoeba induced allergic airway inflammation in mice. We selected 2 free-living amoebas (FLAs) isolated from tap water, which included Korean FLA 5 (KFA5; Vermamoeba vermiformis) and 21 (an homolog of Acanthamoeba lugdunensis KA/ E2). We axenically cultured KFA5 and KFA21. We applied approximately 1 × 106 to mice’s nasal passages 6 times and investigated their pathogenicity. The airway resistance value was significantly increased after KFA5 and KFA21 treatments. The eosinophil recruitment and goblet cell hyperplasia were concomitantly observed in bronchial alveolar lavage (BAL) fluid and lung tissue in mice infected with KFA5 and KFA21. These infections also activated the Th2-related interleukin 25, thymic stromal lymphopoietin, and thymus and activation-regulated chemokines gene expression in mouse lung epithelial cells. The CD4+ interleukin 4+ cell population was increased in the lung, and the secretion of Th2-, Th17-, and Th1-associated cytokines were upregulated during KFA5 and KFA21 infection in the spleen, lung-draining lymph nodes, and BAL fluid. The pathogenicity (allergenicity) of KFA5 and KFA21 might not have drastically changed during the long-term in vitro culture. Our results suggested that Vermamoeba could elicit allergic airway inflammation and may be an airway allergen.