Objective To study the persistence and replication of hepatitis C virus (HCV) RNA in Epstein Barr virus (EBV) transforming peripheral blood mononuclear cell (PBMC) from patients with he patitis C as well as variation of hypervariable regions (HVR) of the HCV genome. Methods PBMC from one patient with hepatitis C was infected by EBV and then transformed into lymphoblasts capable of being propagated indefinitely. Then, HCV RNA of the cultured cells and supernatants was detected by reverse transcriptase polymerase chain reaction (RT PCR) every month and in situ PCR to identify the location of HCV RNA in the cells. The HVR genome sequences of HCV in the first and the ninth month subculture cells were identified by sequence analysis. Results HCV plus strand RNA could be detected in the cultured cells for as long as 1 year. The HCV plus strand RNA could be identified in supernatants and the minus strand RNA were also observed in the cultured cells intermittently. In situ PCR showed that the blue black positive signals of HCV RNA located mainly in cytoplasmas of EBV transforming B cell but negative in nucleous. The positive signal was not found in negative control cells by in situ RT PCR. HCV HVR genome sequence after half year was found to have two regions of a high degree of variability in 1491 to 1583 nucleotide (384 to 414 amino acid) and 1761～1781 nucleotide (473 to 479 amino acid). But, compared the HCV HVR genome sequence in first month subcultured cells with that in ninth month, there was not significant difference. Conclusion HCV may exist in the cultured cell line for a prolonged period wihtout HVR genome sequence changed.

OBJECTIVE To study the immunomodulatory effect of Guipi pills on D-galactose（D-gal）-induced aging mice. METHODS The immune-related targets and related pathways for Guipi pills to exert immune effects were screened by network pharmacology and verified through pharmacodynamic experiments. Totally 105 male ICR mice were randomly divided into blank control （CON） group， model control （MOD） group， positive control （POS） group， Guipi pills low-dose （GD） group and Guipi pills high-dose （GG） group. Except for the CON group， other groups were subcutaneously injected with 400 mg/kg D-gal to induce the aging model； CON group and MOD group were given distilled water， POS group was given 300 mg/kg pidotimod oral solution intragastrically， GD group and GG group were given Guipi pills 300， 600 mg/kg intragastrically， once a day， for 8 weeks. After medication， the serum and spleen were collected， and the contents of interleukin 2 （IL-2）， IL-4， IL-6 and tumor necrosis factor α （TNF-α）， and the contents of immunoglobulin G （IgG）， IgM and IgA were detected. The spleen index was calculated and the histopathological changes in the spleen were observed. The activities of superoxide dismutase （SOD）， glutathione peroxidase （GSH- Px） and malondialdehyde （MDA）， and the content of 8-hydroxy-2 deoxyguanosine （8-OHdG） in spleen were detected； the expression of TNF/phosphoinositide-3-kinase-threonine protein kinase （PI3k-Akt）-related proteins in spleen was detected except for POS group. RESULTS The results of network pharmacology showed that TNF， IL-6 and Akt1 were core targets. The results of pharmacodynamic study showed that compared with MOD group， the contents of IL-2， IL-4， IgG， IgM and IgA were increased significantly in Guipi pills groups， while the contents of TNF-α and IL-6 were decreased significantly； the spleen index was increased significantly （P＜0.05 or P＜0.01）. The phenomenon of diffuse proliferation of lymphocytes was improved， the spleen cells were closely arranged， and the line between the white pulp and red pulp was clear. The activities of SOD and GSH-Px in spleen were increased significantly， while the activity of MDA， the content of 8-OHdG， and the protein expressions of TNF-α， PI3K and p-Akt were decreased significantly （P＜0.05 or P＜0.01）. CONCLUSIONS Guipi pills can regulate the immune function of D-gal-induced aging mice， which is related to regulating the TNF/PI3k-Akt pathway， thereby reducing oxidative stress damage in spleen tissue of mice， and regulating protein expressions of TNF-α， PI3K and p-Akt.

Objective: To explore the expression of CC chemokine receptor 6 (CCR6) in human adenoid cystic carcinoma tissue of salivary glands, and to clarify the mechanism of CCR6 in the nerve invasion of human adenoid cystic carcinoma. Methods: Immumohistochemical method (SP) was performed to detect the expression of CCR6 in 30 cases of cancer tissue of the patients with adenoid cystic carcinoma (including 18 cases in nerve invasion group and 12 cases in non-nerve invasion group) and 22 cases of normal salivary gland tissue. The relationship between the positive expression rate of CCR6 and the clinicopathological features of the patients were analyzed. Results: CCR6 was positively expressed in 22 cases among 30 cases of salivary adenoid cystic carcinoma tissue, and the positive expression rate was 73. 33%. The immunohistochemical staining showed that CCR6 mainly expressed in the cell membrane and cytoplasm of cancer cells. All of the 22 cases of normal salivary gland tissue were negative to CCR6 expression; the difference of the positive expression rate of CCR6 between two groups was statistically significant (P<0. 05). The positive expression rate in nerve invasion group and non-nerve invasion group were 94.44% (17/18) and 41.67% (5/12), respectively; there was statistically significant between two groups (P<0. 05). The positive rates of CCR6 in the patients with salivary gland adenoid cystic carcinoma with different ages, sex and pathological types had no statistically significant differences (P>0. 05). Conclusion: CCR6 plays an important role in the development of human salivary adenoid cystic carcinoma and is closely related to the nerve invasion.

We carried out the stress relaxation and creep experiments on the aorta ascendens of five fresh dead bodies of normal chinese men who had died from impetuous trauma on the head. The stress relaxation and creep data and curves of the aorta ascendens were obtained. Then regression analysis was performed to get the reduced stress relaxation and creep functions. Finally, we analyzed and discussed the results of the experiment.
Adult ; Aorta ; physiology ; Elasticity ; Humans ; In Vitro Techniques ; Male ; Regression Analysis ; Stress, Mechanical ; Viscosity

Objective: To compare the scores of periocular wrinkle and blepharochalasis of the patients before and after the treatment of ultra-pulsed fractional CO2laser with low energy,and to illuminate the efficacy of ultra-pulsed fractional CO2laser with lower energy in eyelid rejuvenation including periocular wrinkle and blepharochalasis.Methods:Thirty patients with periocular wrinkles and blepharochalasis who met the inclusion criteria were selected. The patients were treated with ultra-pulsed fractional CO2laser; the laser parameters were 10 mm spot size, pulse energy 5 mJ and repetition rate 600 Hz and the wrinkle walking areas were treated with 2 mm spot size and the same energy again. Two senior dermatologists scored according to the eyelid laxity scale before treatment and 7 d, 1 month, 3 months and 6 months after treatment, and statistical analysis was performed.Results: Compared with before treatment,there was no significant difference in the scores of eyelid laxity scale 7 d after treatment(P>0.05); the score of eyelid laxity scale was significantly decreased 1 month after treatment (P<0.05);the scores of eyelid laxity scale were significantly decreased 3-6 months after treatment (P<0.01),and the curative effect was more obvious.In this group of 30 patients, there were usually transient erythema, swelling and mild burning pain after treatment. But no serious complications occurred. Conclusion: Ultra-pulsed fractional CO2laser with lower energy(5 mJ) can improve the periocular wrinkles and blepharochalasis effectively with high security and without severe side effects.

Objective: To investigate the tracing fect of new nano-carbon (Kanalin) on the pelvic lymph nodes of the patents with bladder cancer during the laparoscopic radcal cystectomy (RC), and to explain the signcance of its applicaton in the precision surgical treatment. Methods: A total of 44 patents with bladder cancer who underwent RC were divded into tracer group (n=22) and control group (n=22) according to whether Kanalin was used before operaton or not. The laparoscopic RC combined with standard pelvc lymph node dissecon (LND) was performed in the patents in two groups. The effectiveness evaluaton indexes and safety evaluaton indexes of the patents in two groups were collected and compared. The efectiveness evaluaton indexes included black staining situaton of lymph nodes during operaon, the number of detected lymph nodes, the black staining rate of lymph nodes, the number of positive lymph nodes, and operaton time. The safety evaluaton indexes included the blood loss and the indences of perioperative complicatons. Results: The black sainng fect of lymph nodes of the patents in tracer group was definite and there were no tracer-related sde effects and complicatons during the perioperative period. Compared with control group, the clearance rate and detecon rate of lymph nodes of the patents in tracer group were increased (P<0. 01). The black staning rate of lymph nodes of each case was higher than 50%, even up to 85%. There were no significant dfferences in the number of positive lymph nodes, the operaon time, the blood loss and the perioperative complicaton rates of the patents between two groups (P> 0. 05). Conclusion: During the laparoscopic RC, the applicaton of Kanalin for pelvic lymph node tracng is safe, without toxic and side effects; Kanalin is helpful for ncreasing the intraoperative clearance rates and postoperative pathological detecon rates of lymph nodes.

Objective: To compare the changes of soft and hard tissue profiles of the adolescent patients with class II division 1 malocclusion between before and after treated with two different kinds of tooth extraction methods, and to provide the reference for clinical treatment. Methods: A total of 60 adolescent patients with class II division 1 malocclusion were included, and they were divided into 44/44 group (n=30) and 44/55 group (n= 30) based on different kinds of tooth extraction methods. Four first premolars of the patients in 44/44 group were extracted and 2 maxillary first premolars and 2 mandibular second premolars of the patients in 44/55 group were extracted. The measurement indexes of pre-treatment (Tl) and post-treatment (T2) and their difference values (T2 -Tl) of soft and hard tissues of the adolescent patients were detected by lateral cephalometric radiographs. Results: As for the hard tissue, compared with before treatment, the Overjet, SNA, ANB, Ul-NA, distance of Ul-NA, Ul-SN, NA-PA, and A-NP of the adolescent patients in two groups after treatment were decreasd (P< 0. 01), and Ul-Ll and Pog-NB were increased (P<0. 05 or P<0. 01); as for the soft tissue, compared with before treatment, the H angle, UL-SnPg', LL-SnPg', UL-ELine, LL-ELine, and Sn-HLine of the adolescent patients in two groups after treatment were decreased (P<0. 05 or P<0.01); B' LL-FH was increased (P<0. 01); there were no significant differences in the difference values (T2 - Tl) of measurement indexes in the soft and hard tissues of the adolescent patients in two groups (P>0. 05). Conclusion: The changes of soft and hard tissue profiles of the adolescent patients after treated with two different kinds of tooth extraction methods are not significant.

OBJECTIVETo investigate the expression of retinol-binding protein (RBP) in patients with acute myeloid leukemia (AML) and its related factors.

METHODSThe clinical data of 123 patients with AML from October 2012 to February 2016 and 100 healthy controls were collected and the correlation of RBP expression level with sex, French-American-British (FAB) subtypes, molecular mutations, peripheral white blood cells and long-term outcomes was analyzed.

RESULTSCompared with AML patients, the RBP expression level was significantly higher in healthy controls (61.02±34.97 vs 34.44±14.08 mg/L)(u=8.658, P<0.01). Patients with M3 showed higher RBP level (40.74±15.79 mg/L) compared with that of M4 (28.40±13.64 mg/L)(P<0.01) and M5 (31.97±15.31 mg/L)(P<0.05). Negative correlation was observed for RBP and white blood cells in patients with AML (r=-0.352, P<0.001), which was more remarkable in patients with M4 (r=-0.563, P<0.01) and M5 (r=-0.423, P<0.01). AML patients achieved complete remission (CR) (48.64±9.24 mg/L) showed higher RBP level than that before treatment (u=4.876, P<0.01), but lower than healthy controls (u=2.106,P<0.05). After CR, patients with M3 showed higher RBP level than that of non-M3 patients (54.91±5.25 mg/L vs 41.36±7.33 mg/L)(t=8.777,P<0.01).

CONCLUSIONDetecting RBP expression may provide some useful information for urderstanding the pathophysiology and improving the treatment of patients with AML.

BACKGROUND: Circulating tumor DNA (ctDNA) is a promising biomarker for non-invasive epidermal growth factor receptor mutations (EGFRm) detection in lung cancer patients, but existing methods have limitations in sensitivity and availability. In this study, we used the ΔCt value (mutant cycle threshold [Ct] value-internal control Ct value) generated during the polymerase chain reaction (PCR) assay to convert super-amplification-refractory mutation system (superARMS) from a qualitative method to a semi-quantitative method named reformed-superARMS (R-superARMS), and evaluated its performance in detecting EGFRm in plasma ctDNA in patients with advanced lung adenocarcinoma. METHODS: A total of 41 pairs of tissues and plasma samples were obtained from lung adenocarcinoma patients who had known EGFRm in tumor tissue and were previously untreated. EGFRm in ctDNA was identified by using superARMS. Through making use of ΔCt value generated during the detection process of superARMS, we indirectly transform this qualitative detection method into a semi-quantitative PCR detection method, named R-superARMS. Both qualitative and quantitative analyses of the data were performed. Kaplan-Meier analysis was performed to estimate the progression-free survival (PFS) and overall survival (OS). Fisher exact test was used for categorical variables. RESULTS: The concordance rate of EGFRm in tumor tissues and matched plasma samples was 68.3% (28/41). At baseline, EGFRm-positive patients were divided into two groups according to the cut-off ΔCt value of EGFRm set at 8.11. A significant difference in the median OS (mOS) between the two groups was observed (EGFRm ΔCt ≤8.11 vs. >8.11: not reached vs. 11.0 months; log-rank P = 0.024). Patients were divided into mutation clearance (MC) group and mutation incomplete clearance (MIC) group according to whether the ΔCt value of EGFRm test turned negative after 1 month of treatment. We found that there was also a significant difference in mOS (not reached vs. 10.4 months; log-rank P = 0.021) between MC group and MIC group. Although there was no significant difference in PFS between the two groups, the two curves were separated and the PFS of MC group tended to be higher than the MIC group (not reached vs. 27.5 months; log-rank P = 0.088). Furthermore, EGFRm-positive patients were divided into two groups according to the cut-off of the changes in ΔCt value of EGFRm after 1 month of treatment, which was set at 4.89. A significant difference in the mOS between the two groups was observed (change value of ΔCt >4.89 vs. ≤4.89: not reached vs. 11.0 months; log-rank P = 0.014). CONCLUSIONS Detecting EGFRm in ctDNA using R-superARMS can identify patients who are more likely sensitive to targeted therapy, reflect the molecular load of patients, and predict the therapeutic efficacy and clinical outcomes of patients.
Adenocarcinoma of Lung/genetics* ; Circulating Tumor DNA/genetics* ; ErbB Receptors/genetics* ; Humans ; Lung Neoplasms/genetics* ; Mutation/genetics* ; Protein Kinase Inhibitors

Objective: To investigate the effect of enteral nutrition feeding procedure in ICU in severe patients in order to reduce complications and promote rehabilitation of patients. Methods: A total of 184 severe patients admitted to ICU from January to October 2018 were selected as the subjects of the study. 90 patients admitted to the family from January to May were the control group, and 94 patients admitted to the family from June to October were the observation group. The control group adopted routine nutritional support and nursing, and the observation group carried out intensive enteral nutrition feeding process. The nutritional status, immunity, gastrointestinal dysfunction, complications of nosocomial infection and outcome of patients in two groups were compared between the two groups. Results: The nutritional indicators of hemoglobin, serum total protein and serum albumin after a week of treatment were (119.73 ± 9.96), (58.88 ± 2.65), (29.09 ± 1.42) g/L, immunization indicators IgA, IgG, IgM were (2.56±0.10), (2.98±0.36), (1.65±0.15) g/L in the observation group. The control groups were (108.02±9.21), (51.90±2.74), (27.80±1.59), (2.09±0.18), (2.01±0.24), (1.41±0.13)g/L, the difference between the two groups was statistically significant (t=5.81-22.48, P<0.01). The incidence of gastrointestinal dysfunction (gastric retention, diarrhea) in the observation group were 11.70%(11/94), 8.51%(8/94), the incidence of nosocomial infections (ventilator associated pneumonia, catheter related blood stream infection, catheter-associated urinary tract infections) were 6.12‰(5/817), 1.53‰(1/650), 0, the outcome indicators of patients (nutritional standard time, ICU hospitalization time, and mortality rate) were (6.12±1.03) d, (10.98±2.03) d, 13.82%(13/94), all were lower than the control group′s 23.33%(21/90), 20.00%(18/90), 13.06‰(11/842), 11.28‰(8/709), 2.88 ‰(2/694), (7.98 ± 1.54) d, (12.21 ± 1.87) d, 26.67%(24/90), the difference between the two groups was statistically significant (χ²=4.33-8.22, t=6.19, 2.26, P<0.05). Conclusions The procedure of enteral nutrition feeding in patients with severe illness can effectively reduce enteral nutrition complications and enable patients to obtain effective nutrition early.