Objective:To investigate the expressions and clinical significances of histone marks H3K9me3 and H3K27me3 in colorectal cancer.Methods:A retrospective case-control study was conducted. The clinical data of 98 patients with colorectal cancer in Shanxi Province Cancer Hospital from May 2008 to July 2017 were retrospectively analyzed, including 35 patients in the non-metastatic operation-only group, 29 patients in the synchronous hepatic oligometastasis group and 34 patients in the extensive metastasis group, and 33 patients with benign colorectal lesions who underwent colonoscopy in 2017 were selected as the control group. Immunohistochemical assay was used to detect the expressions of H3K9me3 and H3K27me3 proteins in each group, and the expressions of H3K9me3 and H3K27me3 proteins in colorectal cancer patients with different clinicopathological features were analyzed. Kaplan-Meier method was used for survival analysis and log-rank test was performed.Results:The positive expression rate of H3K9me3 protein in colorectal cancer group was 11.2% (11/98), which was lower than that in control group [60.6% (22/33)] ( χ2 = 33.33, P < 0.001); the positive expression rate of H3K27me3 protein in colorectal cancer group was 10.6% (13/98), which was lower than that in control group [97.0% (32/33)] ( χ2 = 76.70, P < 0.001). The positive expression rates of H3K9me3 protein were 60.6% (20/33), 17.1% (6/35), 10.3% (3/29) and 5.9 % (2/34) in the control group, the non-metastatic operation-only group, the synchronous hepatic oligometastasis group and the extensive metastasis group, respectively, and the difference was statistically significant ( χ2 = 26.10, P < 0.001); the positive expression rates of H3K27me3 protein were 97.0% (32/33), 14.3% (5/35), 20.7% (6/29) and 5.9% (2/34), respectively, and the difference was statistically significant ( χ2 = 44.16, P < 0.001). The positive expression rate of H3K27me3 in colorectal cancer tissues of patients with lymph node metastasis degree ≤0.2 was higher than that of patients with lymph node metastasis degree >0.2 [22.4% (11/49) vs. 4.2% (2/48), χ2 = 6.98, P = 0.008]. The median overall survival (OS) time of H3K9me3 positive and negative colorectal cancer patients was 77.0 months (95% CI: 10.6-143.3 months) and 34.0 months (95% CI: 25.5-42.5 months), respectively, and there was no significant difference in OS between the two groups ( P = 0.078). The median OS time of H3K27me3 positive and negative colorectal cancer patients was 39.0 months (95% CI: 15.3- 62.7 months) and 34.0 months (95% CI: 24.3-43.7 months), respectively, and there was no significant difference in OS between the two groups ( P = 0.524). Conclusions:The expressions of H3K9me3 and H3K27me3 in colorectal cancer tissues are lower than those in colorectal benign lesions, and gradually decrease with occurrence of liver metastasis and extensive metastasis. H3K9me3 and H3K27me3 may be potential cancer suppressor factors.

ObjectiveTo investigate the effects of thyme herbal tea （BLX） on the proliferation of human coronavirus OC43 （HCoV-OC43） in vitro and in vivo. MethodThe chemical composition of BLX was analyzed by UPLC-MS. The cytotoxicity of BLX in HRT-18 cells and the effect of BLX treatment on the proliferation of HCoV-OC43 in cells were analyzed. Copies of viral gene were detected by real-time PCR. The effect of BLX treatment on the life cycle of HCoV-OC43 was detected by time-of-addition assay. The maximum tolerated dose of BLX and the influences of BLX on the body weight and survival time of suckling mice infected with HCoV-OC43 were determined. The expression of viral protein in the brain and lung tissue was analyzed by immunohistochemistry. ResultThere were 11 chemical components identified in BLX by UPLC-MS. BLX showed the 50% cytotoxic concentration （CC₅₀） of （13 859.56±319） mg·L^-1， the median inhibitory concentration （IC₅₀） of （1 439.09±200） mg·L^-1， and the selection index of 8.26-11.44 for HCoV-OC43 in HRT-18 cells. Compared with the cells infected with HCoV-OC43， BLX at the concentrations of 1 500， 1 000， 500 mg·L^-1 inhibited the proliferation of this virus （P<0.05， P<0.01）. BLX exhibited antiviral effect in the early stage of virus infection， and the inhibition role in the attachment stage was more significant than that in the entry stage （P<0.05）. In the suckling mice infected with HCoV-OC43， BLX at 1200 and 600 mg·kg^-1·d^-1 alleviated the symptoms， prolonged the survival period， reduced the death rate， and down-regulated the mRNA level of nucleocapsid protein in the mice. Moreover， BLX at 1 200 mg·kg^-1·d^-1 down-regulated the expression of nucleocapsid protein in the brain （P<0.01） and the lung （P<0.01）. ConclusionBLX contained multiple antiviral ingredients. It inhibited the proliferation of HCoV-OC43 both in vitro and in vivo by interference with viral attachment. This study provides theoretical reference for the treatment of acute respiratory tract infection with HCoV-OC43 and for further development and application of BLX.

Objective:To prepare the whole bladder acellular matrix (BAM) using the self-designed perfusion decellularization system, and evaluate the feasibility of constructing the tissue engineering bladder with the adipose-derived stem cells (ADSCs).Methods:This study was conducted from October 2020 to April 2021. The self-designed perfusion decellularization system was used, and four different decellularization protocols (group A, group B, group C and group D) were formulated, according to the flow direction of the perfusate and the action time of different decellularization solutions. Among them, the urethral orifice of the bladder tissue was used as the outflow tract of the perfusion fluid in groups A and B. The top of the bladder was cut off and used as the outflow tract of the perfusion fluid in groups C and D. In groups A and C, 1% Triton X-100 was treated for 6 h, and 1% sodium dodecyl sulfate (SDS) was treated for 2 h. In groups B and D, 1% Triton X-100 was treated for 7 h, and 1% sodium dodecyl sulfate (SDS) was treated for 1 h. In addition, the tissue in the normal bladder group was directly obtained from the natural bladder tissue, which did not require perfusion, cryopreservation and thawing. The fast and efficient decellularization protocol was screened out through HE, DAPI, Masson trichrome and Alcian Blue staining and quantitative analyses to prepare the whole bladder scaffold. The prepared BAM was used as the scaffold material, and the ADSCs were used as the seeding cells to construct the tissue engineering bladder. HE and DAPI staining were used to observe the distribution of ADSCs on the BAM.Results:HE and DAPI staining showed that there was no obvious nuclear residue in the group C. Masson trichrome and Alcian Blue staining showed that the collagen structure and glycosaminoglycan were well preserved in the group C. There was no significant difference in bladder wall thickness between the group C and the normal bladder group [(975.44±158.62)μm vs.(1 064.49±168.52)μm, P > 0.05]. The DNA content in the group C [(43.59 ±4.59) ng/mg] was lower than that in the normal bladder group, group A, group B and group D [(532.50±26.69), (135.17±6.99), (182.49±13.69) and(84.00±4.38)ng/mg], and the difference was statistically significant ( P<0.05). The collagen content [(10.98 ± 0.29)μg/mg] and glycosaminoglycan content [(2.30±0.18)μg/mg] in group C were not significantly different with those in the normal bladder group [(11.69±0.49) and (2.36±0.09)μg/mg, P>0.05]. Scanning electron microscopy showed that a large number of pore structures could be observed on the surface of the prepared BAM in groups A-D and were facilitated to cell adhesion. The isolated and cultured ADSCs were identified by flow cytometry to confirm the positive expression of CD90 and CD29, and the negative expression of CD45 and CD106. Live/dead staining and CCK-8 detection confirmed that the prepared BAM in the group C had no cytotoxicity. HE and DAPI staining showed that a large number of ADSCs were distributed on the surface and inside of the tissue engineering bladder. Conclusions:The whole bladder shape BAM prepared by the self-designed perfusion decellularization system could be used as the scaffold material for bladder tissue engineering, and the constructed tissue engineering bladder could be used for bladder repair and reconstruction.

Objective:This study aimed to investigate the physical properties, biocompatibility, and 3D printing performance of a novel hybrid bioink composed of gelatin methacrylated (GelMA) and chitin nanocrystal (ChiNC).Methods:The study was conducted from May 2021 to December 2022, four different bioinks were prepared by adding varying amounts of ChiNC to GelMA bioink. The GelMA concentration in all four bioinks was 100 mg/ml, while the ChiNC concentrations were 0 mg/ml (no ChiNC added), 5 mg/ml, 10 mg/ml, and 20 mg/ml, respectively, named as GC0, GC5, GC10, and GC20 bioinks. The cross-sectional morphology of the hydrogels formed after photocuring the four bioinks was observed using scanning electron microscopy, and the porosity was calculated. Weighing the hydrogels before and after swelling, and then calculate the equilibrium swelling rate. HUVECs were seeded on the surfaces of the hydrogels prepared from the four bioinks and cultured in medium. Cell proliferation was assessed using CCK-8 assays at 1d, 3d, and 7d to compare the proliferation rates of cells on the four hydrogels. HUVECs were added to the four bioinks, and grid-like scaffolds were printed and cultured in medium. Live-Dead staining was performed at 1d and 7d to observe cell viability. Compare the printing effect of bioinks by observing its forming continuous threads properties during extrusion. Finally, tissue-engineered bladder patches simulating the mucosal layer, submucosal layer, and muscular layer anatomical structures of the bladder wall were 3D bioprinted using the optimized bioink composition, and the stability and fidelity of the printed structures were observed to further validate the feasibility of printing multi-layered complex structures with the bioink.Results:Scanning electron microscopy revealed that the porosity of the GC0, GC5, GC10, and GC20 hydrogels were (51.43±6.23)%, (51.85±6.47)%, (50.55±4.59)%, and (42.49±2.20)%, respectively. The differences in porosity between the GC0 group and the other three groups were not statistically significant ( P=0.9994, P=0.9948, P=0.1200). The equilibrium swelling ratio of the other three groups [(8.81±0.41), (7.95±0.19), (7.71±0.14)] was significantly lower than that of the GC0 group (9.37 ± 0.49), and the differences were statistically significant ( P=0.0457, P<0.01, P<0.01). CCK-8 assay showed no significant difference in absorbance value between the GC10 group (0.360±0.009) and the GC0 group (0.357±0.007), GC5 group (0.350±0.012), and GC20 group (0.345±0.018) on the first day ( P=0.9332, P=0.5464, P=0.4937). However, on the third day, the absorbance value of the GC10 group (0.755±0.012) was significantly higher than that of the GC0 group (0.634±0.010), GC5 group (0.704±0.009), and GC20 group (0.653±0.015) ( P<0.01, P=0.0033, P=0.0002). On the seventh day, the absorbance value of the GC10 group (1.001±0.031) was significantly higher than that of the GC0 group (0.846±0.026), GC5 group (0.930±0.043), and GC20 group (0.841±0.024)( P=0.0012, P=0.1390, P=0.0010). The addition of human umbilical vein endothelial cells (HUVECs) into the four groups of hydrogels enabled the printing of grid-like scaffolds, and Live-Dead staining was performed on day 1 and day 7. The cell viability of HUVECs in the four groups on day 1 was (90.13±1.63)%, (90.6±2.45)%, (92.58±2.15)%, and (91.40±3.17)%, respectively. There were no statistically significant differences between the GC0 group and the other three groups ( P=0.9869, P=0.3093, P=0.8008). On day 7, the cell viability was (89.97±3.10)%, (92.18±2.21)%, (92.05±2.25)%, and (90.12±1.97)% for the four groups, respectively. There were no statistically significant differences between the GC0 group and the other three groups ( P=0.3965, P=0.4511, P=0.9995). Bioink extrusion test showed that the GC0 hydrogel could be extruded continuously and form threads at temperatures between 24℃ and 25℃, while the GC10 hydrogel could be extruded continuously and form threads at temperatures between 24℃ and 27℃. Printing tissue engineered bladder patches simulating the anatomical structure of the bladder mucosal layer, submucosal layer, and muscular layer using GC10 bioink, and the printed patches were stable, without collapse, and had high fidelity. Conclusions:Adding ChiNC to GelMA promotes cell adhesion, proliferation, and expands the printing window of GelMA bioink. The biocompatibility of the mixed bioink prepared by adding 10 mg/ml ChiNC in GelMA is good, capable of printing tissue-engineered bladder patches that mimic the anatomical structure of natural bladder walls.

Objective:To investigate the effect of tissue engineered bladder patch constructed by double-layer silk scaffold and adipose-derived stem cells (ADSCs) in the repair and reconstruction of bladder.Methods:This study was conducted from May 2020 to March 2021. The silk fibroin (SF) aqueous solution was obtained from silkworm cocoons, and a double-layer silk scaffold composed of silk fibroin film and silk fibroin sponge was further prepared. The rat ADSCs were isolated, cultured, and the ADSCs surface markers (CD29, CD90, CD45, CD106) were identified by flow cytometry. The ADSCs were planted on a double-layer silk scaffold to construct a tissue-engineered bladder patch. Thirty-six male SD rats were randomly divided into three groups: tissue engineered bladder patch group (SF-ADSCs group, n=15), double-layer silk scaffold group (SF group, n=15), control group ( n=6). The tissue engineered bladder patch (SF-ADSCs group) and double-layer silk scaffold (SF group) were wrapped on the omentum to promote vascularization. The vascularization was evaluated by HE and immunofluorescence staining. The wrapped tissue engineered bladder patch and double-layer silk scaffold were used to repair the defective bladder. In the control group (six rats), the incision was closed immediately after the bladder tissue fully exposed. At 4 weeks and 12 weeks after operation, the general morphology of bladder tissue and cystography were performed to evaluate the recovery of bladder morphology. After the graft was harvested, HE and Masson's trichrome staining and immunofluorescence staining were used to observe the regeneration of bladder wall tissue. Urodynamics was used to assess the recovery of bladder function at 12 weeks after operation. Results:The flow cytometry results confirmed that the isolated cells positively expressed CD29 and CD90, and there was no significant expression of CD45 and CD106. Gross observation and scanning electron microscope confirmed that the preparation of double-layer silk scaffold not only had a pore structure that was conducive to cell planting, but also had good toughness and was facilitated to surgical suture. The number (43.50±2.66) and area (0.73±0.03)% of vascular-like structures in the SF-ADSCs group after the omentum encapsulation was significantly higher than that in the SF group [(24.50±3.51), (0.55±0.05)%], and the difference was statistically significant ( P<0.05). At 4 weeks after bladder repair, the histological staining of the grafts in the SF-ADSCs and SF groups showed a large number of degraded fragments of double-layer silk scaffold. At 12 weeks, the morphology of the graft in the SF-ADSCs group showed uniform bladder morphology, which was similar to that of normal bladder tissue. Immunofluorescence staining showed that the continuous urothelial layer, abundant smooth muscle tissue, vascular structure and regenerated neurons could be observed in the SF-ADSCs group. Urodynamic test showed that the bladder maximum volume (0.74±0.03)ml and compliance (16.68±0.44)μl/cm H 2O in the SF-ADSCs group, which were better than that in the SF group [(0.47±0.05)ml, (14.89±0.37)μl/cm H 2O], but lower than that in the control group [(1.12±0.08)ml, (19.34±0.45)μl/cm H 2O], and the difference was statistically significant ( P<0.05). Conclusions:The tissue engineered bladder patch constructed with double-layer silk scaffolds and ADSCs could promote the morphological repair of bladder tissue, the regeneration of bladder wall structure and the recovery of bladder physiological function.

Background: and Purpose Brainstem gliomas (BSGs) in adults are rare brain tumors with dismal outcomes. The aim of this study was to determine the clinical and genetic features in a series of BSGs and their association with the prognosis. Methods: Fifty patients who underwent a stereotactic biopsy between January 2016 and April 2018 at a single institution were collected. Data on clinicopathological characteristics were analyzed and factors associated with patient survival were identified using a Cox regression model. Results: The median age at diagnosis was 55.5 years, and 62% of the patients were male. Glioblastoma (44%) accounted for the largest proportion of BSGs, and oligodendroglioma (2 of 50) was rarely encountered. The IDH mutation (6 of 44) occurred infrequently in astrocytomas, and IDH-mutant tumors harbored both ATRX loss and MGMT promoter methylation at a relatively low level. Wild-type IDH astrocytomas were identified as having high rates of 1p/19q codeletion (5 of 38) and loss of heterozygosity 1p (8 of 38) or 19q (8 of 38) only. In diffuse midline glioma H3K27M mutant, MGMT promoter methylation occurred in three of four cases. Patients were offered radiotherapy and/or concurrent/adjuvant temozolomide chemotherapy, and their median survival time was 13 months. Multivariate analysis revealed that a low tumor grade, absence of tumor enhancement, duration of symptoms ≥3 months, Karnofsky performance status ≥70, and ATRX loss conferred a survival advantage. Conclusions Adult BSGs showed different molecular genetic characteristics, but also resembled supratentorial gliomas in their clinical features associated with oncological outcomes.

Background: and Purpose Brainstem gliomas (BSGs) in adults are rare brain tumors with dismal outcomes. The aim of this study was to determine the clinical and genetic features in a series of BSGs and their association with the prognosis. Methods: Fifty patients who underwent a stereotactic biopsy between January 2016 and April 2018 at a single institution were collected. Data on clinicopathological characteristics were analyzed and factors associated with patient survival were identified using a Cox regression model. Results: The median age at diagnosis was 55.5 years, and 62% of the patients were male. Glioblastoma (44%) accounted for the largest proportion of BSGs, and oligodendroglioma (2 of 50) was rarely encountered. The IDH mutation (6 of 44) occurred infrequently in astrocytomas, and IDH-mutant tumors harbored both ATRX loss and MGMT promoter methylation at a relatively low level. Wild-type IDH astrocytomas were identified as having high rates of 1p/19q codeletion (5 of 38) and loss of heterozygosity 1p (8 of 38) or 19q (8 of 38) only. In diffuse midline glioma H3K27M mutant, MGMT promoter methylation occurred in three of four cases. Patients were offered radiotherapy and/or concurrent/adjuvant temozolomide chemotherapy, and their median survival time was 13 months. Multivariate analysis revealed that a low tumor grade, absence of tumor enhancement, duration of symptoms ≥3 months, Karnofsky performance status ≥70, and ATRX loss conferred a survival advantage. Conclusions Adult BSGs showed different molecular genetic characteristics, but also resembled supratentorial gliomas in their clinical features associated with oncological outcomes.

Ulcerative Colitis (UC) has been reported to be related to Porphyromonas gingivalis (P. gingivalis). Porphyromonas gingivalis peptidylarginine deiminase (PPAD), a virulence factor released by P. gingivalis, is known to induce inflammatory responses. To explore the pathological relationships between PPAD and UC, we used homologous recombination technology to construct a P. gingivalis strain in which the PPAD gene was deleted (Δppad) and a Δppad strain in which the PPAD gene was restored (comΔppad). C57BL/6 mice were orally gavaged with saline, P. gingivalis, Δppad, or comΔppad twice a week for the entire 40 days (days 0-40), and then, UC was induced by dextran sodium sulfate (DSS) solution for 10 days (days 31-40). P. gingivalis and comΔppad exacerbated DDS-induced colitis, which was determined by assessing the parameters of colon length, disease activity index, and histological activity index, but Δppad failed to exacerbate DDS-induced colitis. Flow cytometry and ELISA revealed that compared with Δppad, P. gingivalis, and comΔppad increased T helper 17 (Th17) cell numbers and interleukin (IL)-17 production but decreased regulatory T cells (Tregs) numbers and IL-10 production in the spleens of mice with UC. We also cocultured P. gingivalis, Δppad, or comΔppad with T lymphocytes in vitro and found that P. gingivalis and comΔppad significantly increased Th17 cell numbers and decreased Treg cell numbers. Immunofluorescence staining of colon tissue paraffin sections also confirmed these results. The results suggested that P. gingivalis exacerbated the severity of UC in part via PPAD.
Animals ; Colitis, Ulcerative/microbiology* ; Mice ; Mice, Inbred C57BL ; Porphyromonas gingivalis/pathogenicity* ; Protein-Arginine Deiminases ; Virulence Factors

Objective: To investigate the expression level of antisense transcript of pseudogene, general transcription factor Ⅱi psedugen23 (GTF2IP23), in breast cancer and its effect on the host gene general transcription factor Ⅱi (GTF2I). Methods: The expressions of GTF2IP23 and GTF2I were detected in 40 cases of invasive breast cancer tumors and their counterparts by using quantitative real-time polymerase chain reaction (qRT-PCR). The effects of GTF2IP23 on the expression of GTF2I gene and cell proliferation and migration were analyzed by overexpression of GTF2IP23 in breast cancer cells. Results: The expression of GTF2IP23 mRNA in breast cancer tissues was significantly higher than that in adjacent tissues (P<0.001), while the expression of GTF2I mRNA was significantly lower than that in adjacent tissues (P=0.007). The expression of GTF2IP23 was negatively correlated with GTF2I (r=-0.335, P=0.025). The expression of GTF2IP23 in breast cancer cells was significantly higher than in normal breast cells (P<0.01), while GTF2I expression in breast cancer cells was significantly lower than that in normal breast cells (P<0.01). Overexpression of GTF2IP23 in ZR-75-30 cells significantly reduced the expression of GTF2I (P=0.034) and enhanced cell proliferation (P=0.017) and migration (P=0.026) capacity. Conclusions GTF2IP23 is distinctly upregulated in breast cancer, it inhibits the expression of real gene GTF2I and promotes the proliferation of breast cancer cells.

Objective To investigate the effect of different enteral nutrition (EN) preparations on nutritional status and glucose metabolism in elderly heart failure (HF) patients. Methods Eighty-two elderly HF patients were consecutively admitted to Taizhou Enze Medical Center Group Enze Hospital from January 2017 to March 2018, and they were randomly divided into a study group and a control group, each group with 41 cases. The basic treatments were the same in two groups, while the patients in study group were treated with TPF-D emulsion, and those of control group were treated with SP emulsion. Nutritional status [serum albumin (Alb), proalbumin (PA) and transferrin (TF)], glucose metabolism indexes (glucose, Betatrophin and insulin dose), cardiac function [N-terminal B-type pro-brain natriuretic peptide (NT-proBNP), left ventricular ejection fraction (LVEF)], high-sensitivity C-reactive protein (hs-CRP) in serum were compared between two groups after treatment, and the incidences of adverse reactions (gastric retention, diarrhea, constipation, electrolyte abnormality, pulmonary infection, dysfunction of liver) were also observed in the two groups. Results In the aspect of nutrition, no statistical significant differences in indexes were found at each time point between the two groups during the therapeutic course (all P > 0.05). In terms of glucose metabolism, the blood glucose levels in the study group were significantly lower than those in the control group on day 5 and day 7 after treatment (mmol/L: 9.73±3.69 vs. 11.56±3.78 on day 5, 8.35±2.17 vs. 10.41±2.38 on day 7, both P < 0.05); the serum Betatrophin levels in the study group were also significantly lower than those in the control group on 5 days and 7 days (μg/L: 5 days was 1.36±0.49 vs. 1.89±0.74, 7 days was 1.31±0.47 vs. 1.62±0.59, both P < 0.05); the doses of insulin used in the study group were obviously lower than those in the control group on 3, 5 and 7 days (U: 3 days was 37.86±8.49 vs. 47.84±10.33, 5 days was 29.33±7.45 vs. 38.56±9.69, 7 days was 13.35±8.56 vs. 28.49±9.77, all P < 0.05). In terms of cardiac function, the study group showed significant lower NT-proBNP levels on 3, 5 and 7 days (μg/L: 3 days was 5.8±0.8 vs. 6.2±1.0, 5 days was 3.7±0.7 vs. 4.4±0.9, 7 days was 1.9±0.5 vs. 3.1±0.7, all P <0.05]; significant higher level of LVEF in study group on 7 days (0.50±0.02 vs. 0.48±0.03, P < 0.05) and significant lower hs-CRP levels in study group on 5 days and 7 days (μg/L: 5 days was 27.12±10.58 vs. 34.41±12.69, 7 days was 15.33±9.87 vs. 22.71±11.45, both P < 0.05). In addition, compared with the control group, the incidence of gastric retention [17.07% (7/41) vs. 36.59% (15/41)], so of diarrhea [14.63% (6/41) vs. 34.15% (14/41)] and of electrolyte abnormalities [14.63% (6/41) vs. 39.02% (16/41)] were significantly lower in study group than those in the control group (all P <0.05). Conclusion Compared with SP, TPF-D not only can effectively improve the nutritional status, but also can superiorly control blood glucose, improve cardiac function and lower the incidence of adverse reactions in elderly HF patients.