Objective To establish the quality standard for Shengxinfa Capsule. Methods TLC was used to identify Praeparata, Psoraleae Fructus and Gastrodiae Rhizoma in Shengxinfa Capsule, and 2,3,5,4’-the four-stilbene-2-O-β-D-glucoside was identified by HPLC. The column of Agilent TC-C18 (4.6 mm×250 mm, 5 μm) was used. The mobile phase was acetonitrile-water (15∶85). The flow was 1.0 mL/min and column temperature was 30 ℃. UV detecting wavelength was 320 nm. Results Praeparata, Psoraleae Fructus and Gastrodiae Rhizoma could be identified by TLC. The linear range of 2,3,5,4’-the four-stilbene-2-O-β-D-glucoside was in the range of 0.05-0.50 μg. The average recovery was 97.8% (RSD=0.39%). Conclusion The method is feasible and accurate. The quality of Shengxinfa Capsule can be controlled effectively with the established quality standard.

Objective To investigate the roles of endoscopic biopsy and third ventriculostomy in the diagnosis and treatment of pineal region tumors in children. Methods Endoscopic biopsy and third ventriculostomy were performed in 9 pediatric patients with pineal region tumors. Results Successful third ventriculostomy, confirmed by MRI, was performed in 9 cases of children with obstructive hydrocephalus. No complications were found in all patients. Conclusion Endoscopic biopsy and third ventriculostomy are effective neuroendoscopic procedures in minimally invasive preferential management of pineal region tumors.

AIM: To investigate the effect of silymarin on homocysteine-induced cell viability and apoptosis in human umbilical vein endothelial cells (HUVECs). METHODS: Cell viability was analyzed by using MTT and LDH assay. Apoptotic cells were detected by using DNA fragmentation and flow cytometric analysis. The level of intracellular reactive oxygen species (ROS) and the potential of mitochondrial membrane were determined by flow cytometric assay. The activity of caspase-3, -6 and -9 were measured with microplate spectrofluorometer. Protein levels were examined by Western blotting. RESULTS: Treatment of cultured HUVECs with HCY for 48 h induced a significant decrease in cell viability, and the percentage of apoptosis increased to 76.8%. The level of intracellular ROS and activity of caspase-3, -6 and -9 enhanced, and the red/green ratios of mitochondrial membrane decreased. However, simultaneous treatment with silymarin exhibited cytoprotective effects, reduced formation of the DNA ladder, prevented the levels of Bax and Bcl-2 proteins and the accumulation of ROS as well as caspase-3, -6 and -9 activation, reconverted the potential of mitochondrial membrane, and the percentage of apoptosis/necrosis was significantly decreased to 12.7% in a dose-dependent manner. CONCLUSION: These results demonstrate that silymarin has the protective capacity to antagonize HCY-induced apoptosis in HUVECs. The antiapoptotic action of silymarin may be partially dependent on an anti-oxidative stress effects, inhibition of caspases activity, and maintenance of mitochondria function.

Objective To explore the expression level and significance of the nod-like receptor protein 3 (NLRP3) inflammasome in renal tissue with calcium oxalate stone.Methods 20 kidney specimens were collected as the experimental group from patients with calcium oxalate stone who underwent nephrectomy because of stones in our hospital between January 2008 and December 2014;another 20 renal specimens were get as the control group from patients with renal carcinoma,the renal tissues were obtained 2cm far from the tumor and proved as normal tissue.Immunohistochemical detection was carried out to analyze the expression level of NLRP3,Caspase-1,and IL-1β in the 40 renal samples.Animal experiment:fourteen male SD rats were randomly divided into calcium oxalate stone group and control group.For calcium oxalate stone group we established an ethylene glycol method induced hyperoxaluric rat model featured by crystalline material within tubule lumens;for control group normal feeding was performed.After 6 weeks,all rats were sacrificed,and the kidneys were harvested for further experiments.HE staining and Pizzolato staining were used to detect calcium oxalate crystals within tubule lumens.Western boltting and RT-PCR was applied to detect protein level and mRNA quantity of NLRP3,Caspase-1,and IL-1β from tissue lysates in rat model.Results In renal tissue samples obtained from patients with calcium oxalate stone disease,we demonstrated that the expression level of NLRP3,Caspase-1,and IL-1β were above to the normal renal tissue samples.We established a hyperoxaluric rat model character with crystalline material within tubule lumens examined by renal histology with HE staining and Pizzolato staining.And we detected that the protein and mRNA levels of NLRP3,Caspase-1 and IL-1β were remarkably increased in the lysates from the hyperoxaluric rat model (P ＜ 0.05).Conclusions The NLRP3 inflammasome has overexpression in the renal tissue of patients with calcium oxalate stone as well as in the renal tissue of hyperoxaluric rat,and it provides a new thought to reveal the formation of calcium oxalate stone.

Objective: To discuss the correlation between myeloid derived suppressor cells (MDSCs) and hepatic trans-planted tumor and to explore new ways to inhibit the development of hepatic cancer. Methods: We established the animal models with H_(22) hepatic carcinoma cells transplanted to the anterior right limb. Then the MDSCs morphology was observed with confocal microscopy and the proportion of MDSCs in blood and spleen was measured with flow cytometry. The 36 mice were divided into three groups: the control group, the low-dose group (2mg/kg) and the high-dose group (4mg/kg). Then As_2O_3 was injected twice a week to the mice before repeating the aforementioned measures. The direct effects of As_2O_3 on MDSCs cultured with H_(22)-ascites supernatant was observed. Results: At 25 days after transplantion, the tumor weight was increased to 5.67g, and the proportion of MDSCs in blood and spleen was increased to 20.46% and 9.50%, re-spectively. There was a positive correlation between hepatic transplanted tumor and MDSCs in blood and spleen and the relative factors were 0.95 and 0.96, respectively (t=-5.270 and 5.939, P<0.05). With the effect of As_2O_3, the proportion of MD-SCs in blood in low-dose group and high-dose group was 11.31% and 10.00% at 28 days after treatment, lower than that in the control group (t=3.193 and 5.486, P<0.05), and there was also a statistical difference between the high-dose group and low-dose group (t=3.066, P<0.05). The proportion of MDSCs in the spleen in low-dose group and high-dose group was 10.90% and 9.04% at 28 days, lower than that in the control group (t=3.586.and 5.279, P<0.05), but there was no statistical difference between the high-dose group and low-dose group (1=1.298, P>0.05). In vitro, the proportion of MDSCs in nutrient fluid was increased to 12.67% at 12 days after treatment with H_(22)-ascites supematant, and was decreased to 7.44% at 18 days after treatment with As_2O_3. Conclusion: The proportion of MDSCs in H_(22) tumor-bearing mice is increased because of tu-mor development. There is a positive correlation between MDSCs and hepatic transplanted tumor. As_2O_3 can decrease MD-SCs and inhibit tumor growth.

OBJECTIVETo study how the way in which betaine promotes the proliferation of mouse spleen lymphocytes is related to calcium channels.

METHODBALB/c mice were used for this experiment. Mouse spleen lymphocytes were obtained through in vitro cultivation after they had been separated, and were divided into a negative control group, a Con A group, and 0.04, 0.4, 4, and 20 mmol x L(-1) betaine groups. MTT was used to observe the effect of betaine on the proliferation of mouse spleen lymphocytes; flow cytometry was used to measure the changes in the cell cycle of mouse spleen lymphocytes; and laser confocal scanning microscopy was used to observe the changes in the intracellular [Ca2+]i of mouse spleen lymphocytes after betaine or different calcium channel blockers were applied.

RESULTBetaine was found to promote the proliferation of mouse spleen lymphocytes 12 h after it had been applied in vitro in concentrations of 4 and 20 mmol x L(-1). It was also found to promote the proliferation of mouse spleen lymphocytes 24 h and 48 h after it had been applied in vitro in concentrations of 0.04, 0.4, 4, and 20 mmol x L(-1), with the effect being most marked for the 4 mmol x L(-1) group 24 h after its application. It was found to facilitate the entry of mouse spleen lymphocytes from the G0/G1 to the S phase 4, 6, 18, and 24 h after it had been applied to mouse spleen lymphocytes in a concentration of 4 mmol x L(-1), with the effect being most marked at 18 h after its application. Intracellular [Ca2+]i in mouse spleen lymphocytes increased significantly (P < 0.01) 6, 12, 18 h after 4 mmol x L(-1) betaine had acted on the lymphocytes, with the effect being most marked at 6 h. The calcium channel blockers nifidipine, diltiazem, mibefradil, and genistein had no effect on the increase of the intracellular [Ca2+]i in mouse spleen lymphocytes due to the application of betaine, while verapamil, mycifradin, heparin, and procaine could block such increase.

CONCLUSIONBetaine facilitates the entry of mouse spleen lymphocytes from the G0/G1 into the S phase by raising the intracellular [Ca2+]i in these cells, thus promoting their proliferation. Intracellular [Ca2+]i increases mainly in two ways: (1) By affecting the alpha1 subunit of the L-type voltage-gated calcium channel with mediation by G proteins and thus leading to an efflux of intracellular calcium: (2) By affecting the IP3R and RyR calcium channels of the intracellular calcium stores and thus leading to the release of intracellular calcium.

Animals ; Betaine ; pharmacology ; Calcium ; metabolism ; Calcium Channels ; metabolism ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Female ; Lymphocytes ; cytology ; drug effects ; metabolism ; Male ; Mice ; Mice, Inbred BALB C

BACKGROUND AND OBJECTIVEThe CIK cell is one of the most important means of the adoptive cellular immunotherapy, and it is a hotspot of which to simplify its culture procedure and to promote its inhibition rate. The aim of this study is to observe the biological function of the CIK cells cultivated by the recombinant human fibronectin (RN) and to establish an effective and simple way of cells expansion.

METHODSWe separated the mononuclear cells (PBMCs) in 50 mL peripheral blood from 10 healthy persons with density gradient centrifugation in the lymphocyte-separating medium, and the PBMCs were divided into two groups, of which were cultivated by RN-introduced and conventional method separately. Then we estimated the proliferation ability, and analyzed the immunologic type, IFN-gamma, IL-4, perforin and granzyme B of them with flow cytometry. Besides that, we tested the inhibition rate of CIKs cells to four kinds of human lung cancer cell lines in vitro by MTT assay.

RESULTSThe RN-induced group had a higher proliferation rate that was 2.0-3.5 times of the conventional group, and there was an obvious statistical difference between the two (P < 0.05). The proliferation rates of CD3+CD16+CD56+T cells in each group were 3 778 and 2 068 times of the initial number, respectively. There was also a higher percentage of CD3+CD8+ T cells in RN-induced group (P < 0.05), while the percentage of CD3+CD4+T cells had no significant statistical difference (P > 0.05). We found a similar inhibition rate of the CIK cells to all this human lung cancer cell lines (P > 0.05). The cells which secreted IFN-gamma increased, while the cells which secreted IL-4 did not. The cells which secreted granzyme B and perforin were positive.

CONCLUSIONIt is an effective and simple way to cultivate the CIK cells with RN, which should be adopted.

CD3 Complex ; metabolism ; CD4-Positive T-Lymphocytes ; immunology ; CD56 Antigen ; metabolism ; Cell Line, Tumor ; Cells, Cultured ; Cytokine-Induced Killer Cells ; drug effects ; metabolism ; Fibronectins ; genetics ; metabolism ; pharmacology ; Flow Cytometry ; Humans ; Immunotherapy, Adoptive ; methods ; Interleukin-4 ; metabolism ; Lung Neoplasms ; therapy ; Receptors, IgG ; metabolism ; Recombinant Proteins ; genetics ; metabolism ; pharmacology ; T-Lymphocytes ; immunology

BACKGROUND AND OBJECTIVEThe actual evaluation of immunological function is significant for studing the tumor development and devising a treatment in time. The aim of this study is to evaluate the immunological function of advanced lung cancer patients systematically, and to discuss the clinical significance.

METHODSThe nucleated cell amounts of advanced lung cancer patients and the healthy individuals were counted. The immune cell subsets and the levels of IL-4, INF-gamma, perforin and granzyme in CD8+T cells by the flow cytometry were measured. The proliferation activity and the inhibition ratio of immune cells to several tumor cell lines were evaluated by MTT assay.

RESULTSThe absolute amounts and subsets of T, B, NK cells of advanced lung cancer patients were lower than the healthy individuals (P < 0.05); However, the proportion of regulatory T cells of advanced lung cancer patients (4.00 +/- 1.84)% was lower than the healthy individuals (1.27 +/- 0.78)% (P < 0.05). The positive rates of IFN-gamma perforin, granzyme in CD8+T cells decreased while them in IL-4 did not in the advanced lung cancer patients compared to the healthy control group (P < 0.05). The proliferation activity of immune cells, the positive rate of PPD masculine and the inhibition ratio to tumor cells in the advanced lung cancer patients was lower than the healthy subsets obviously (P < 0.05).

CONCLUSIONThere was a significant immune depression in the advanced lung cancer patients compared to the healthy individuals.

Adult ; Aged ; CD8-Positive T-Lymphocytes ; immunology ; metabolism ; Cell Line, Tumor ; Female ; Flow Cytometry ; Granzymes ; metabolism ; Humans ; Interferon-gamma ; metabolism ; Interleukin-4 ; metabolism ; Lung Neoplasms ; immunology ; pathology ; Male ; Middle Aged ; Perforin ; metabolism

Objective:To observe the clinical and multimodal imaging characteristics of eyes with acute macular neuroretinopathy (AMN) associated with COVID-19.Methods:A retrospective clinical study. From December 18 to 26, 2022, 16 eyes of 8 patients with AMN associated with COVID-19 were included in the study. There were 4 males and 4 females; all cases were bilateral. The age was (31.5±9.6) years old. The time from fever to decreased vision was (3.75±1.04) days. The best corrected visual acuity (BCVA), intraocular pressure, slit lamp microscopy, indirect fundus microscopy, fundus color photography, and optical coherence tomography (OCT) were performed in all patients. Infrared fundus photography (IR), OCT angiography (OCTA) and fluorescein fundus angiography (FFA) were performed in 14, 6 and 4 eyes respectively. The BCVA examination was performed using the international standard visual acuity chart, which was converted into logarithm of the minimum angle of resolution (logMAR) BCVA for statistics. The clinical data, IR, OCT and OCTA imaging features of the patients were retrospectively analyzed.Results:The logMAR BCVA of AMN eyes was 4.21±0.74, intraocular pressure was (14.87±1.50) mm Hg (1 mm Hg=0.133 kPa). Fundus color photography showed that multiple gray-white petal-shaped lesions were arranged around the macular fovea in 2 eyes; no obvious abnormality was found in the macular area in 14 eyes. Of the 14 eyes examined by IR, 6 eyes had irregular weak reflective lesions around the macular fovea. OCT showed strong reflex in the outer nuclear layer and outer plexiform layer of all eyes, including 15 eyes with elliptical zone injury. In 6 eyes examined by OCTA, the blood flow density of the superficial and deep capillary plexus (DCP) of retina decreased, and the blood flow density of DCP decreased significantly. The en-face image of DCP showed the wedge-shaped strong reflective lesion area with the tip pointing to the central fovea in 2 eyes. No abnormal fluorescence was observed in FFA.Conclusions:The characteristic manifestation of AMN associated with COVID-19 is weak reflex focus in IR; OCT shows strong reflection in outer core layer and outer plexiform layer; OCTA showed that retinal DCP blood flow density decreased.

OBJECTIVE：To observe the p rotective effects of resveratrol （Res）on LPS-induced acute lung injury （ALI）model mice，and to explore its possible mechanism based on TLR 4/NF-κB pathway. METHODS：Kunming mice were divided into normal group，model group ，positive control group （dexamethasone，0.5 mg/kg），Res low-dose ，medium-dose and high-dose groups （50， 100，200 mg/kg），with 10 mice in each group. Normal group and model group were given normal saline intragastrically ，once a day，for 7 days；positive control group were intraperitoneally injected with Dexamethasone sodium phosphate injection ，once a day，for 3 days；Res groups were given relevant medicine intragastrically ，once a day ，for 7 days. After last administration ，all the mice except the normal group were dripped LPS （5 mg/kg）into the nose to induce ALI model. The apoptosis of neutrophils in BALF was observed by Hoechst 33242 staining；the apoptosis rate of neutrophils were detected by flow cytometry. The contents of IL- 6 and TNF-α in the plasma were detected by ELISA. After wet mass to dry mass （W/D）ratio of lung was E-mail：492234709@qq.com calculated，the morphological characteristics of lung tissue were observed by HE staining. Western blotting assay was used to detect the expression o f TLR 4 and NF-κ B in lung tissue. RESULTS ：In normal group ，there were few apoptotic neutrophils in the BALF ，and the lung tissue structure was intact ， without edema，hyperemia，exudation，inflammatory cell infiltration or other inflammatory manifestations. In model group ，the number of apoptotic neutrophils in BALF increased ，and the apoptotic rate of neutrophils were enhanced significantly （P＜0.01）； edema and hyperemia of lung tissue were significantly increased ，and the red consolidation area was observed ；the contents of IL- 6 and TNF-α in plasma，the ratio of lung W/D，the relative expression of TLR 4 and NF-κB in lung tissue were significantly increased （P＜0.01）. Compared with model group ，the number of apoptotic neutrophils in BALF were increased ，and the apoptotic rate of neutrophils were enhanced significantly （P＜0.05 or P＜0.01）above symptoms of lung tissue were improved to different extents ； the contents of IL- 6 and TNF-α in plasma，lung W/D ratio as well as relative expression of TLR 4 and NF-κB in lung tissue（except for Res low-dose group ）in administration groups were decreased significantly （P＜0.05 or P＜0.01）. CONCLUSIONS ：Res has a protective effect on ALI model mice ，the mechanism of which may be related to reducing the generation of inflammatory cytokines TNF-α and IL-6 by inhibiting TLR 4/NF-kB expression.