Objective To explore the bioeffect of different parameters on 4 cell lines by ultrasoundmediated microbubble destruction.Methods The orthogonal experimental design was used to investigate the effect of three factors on the bioeffects of four cell lines under three levels.Three factors included microbubble concentration,sound intensity,irradiation time.Human breast tumor (MCF-7) cells,ovarian tumor (A2780) cells,liver tumor (Bel7402) cells and thyroid tumor (ARO) cells were exposed to ultrasound in the presence of SonoVue.The cell survival rate was determined by MTT methods and the cell luminosity factor was detected by flow cytometry.Results The optimum parameters for Bel7402 and ARO cell were the same (A2B3C2),and they were different from those from MCF-7 (A3B1C1) and A2780 (A1B3C3) cell.The cell survival rates for 4 cell lines were above 75％,and the cell luminosity factors were different among 4 cell lines.Conclusions The optimum parameters by ultrasound-mediated microbubble destruction for different cell lines are different,and under the optimum parameters the bioeffects of different cell lines are different.

Objective To investigate the selective targeting ability of a novel folate-modified nanobubbles with two-fold amount of folate [(FOL)2-NBs] . Methods DSPE-PEG2000-AD-(FOL)2with two-fold of folate per DSPE-PEG2000 chain was synthesized and then tested by 1 H nuclear magnetic resonance (1H NMR) . The novel (FOL)2-NBs was prepared using the mechanical shaking method based on lipid-stabilized perfluoropropane . The bubble size was measured by Malvern laser particle size analyzer and the contrast enhancement ability was also detected with imaging machine using a self-made agarose mold . The experiment of selective targeting ability was also carried out in human breast cancer MCF-7 cell with over-expression of folate receptor ( FR) using fluorescence activated cell sorting ( FACS) . Results The result of 1H NMR proved that DSPE-PEG2000-AD-( FOL )2was successfully synthesized ,and the purity reached up to 90% . The novel prepared ( FOL) 2-NBs showed superior contrast enhancement ability with a particle size of ( 286 .87 ± 22 .96) nm . Compared with the conventional NBs ,the novel ( FOL) 2-NBs exhibited improved selective cellular targeting ability proven by FACS . Conclusions A novel nanobubble with improved selective targeting ability is successfully prepared and shows great potential in extravascular imaging and curation in FR overexpressed tumors .

Objective:To explore the clinical characteristics of elderly patients with type 2 diabetes mellitus (T2DM) complicated with coronary atherosclerotic heart disease (CHD) and analyze the risk factors of CHD in patients with T2DM.Methods:Using the method of retrospective cohort study, 406 elderly male patients with T2DM (≥75 years old) admitted to Xinhua Hospital Affiliated to Shanghai Jiaotong University from January 2017 to January 2020 were selected and divided into T2DM without CHD group (165 cases) and T2DM with CHD group (241 cases). The clinical characteristics in elderly patients with T2DM complicated with CHD and risk factors for CHD were analyzed.Results:The age ((86.78±5.35 )years old), course of T2DM((12.32±0.46) years), fasting blood glucose(FPG)((7.64±2.81) mmol/L), hemoglobin a1c (HbA1c)((7.59±1.21)%), the proportion of hypertension(84.65%(204/241)), D-dimer((0.50±0.13) mg/L), the incidence of thromboembolic events(46.06%(111/241)), blood serum creatinine ((94.81±12.70) μmol/L), urea nitrogen((8.31±4.46) mmol/L), uric acid((376.44±116.01) μmol/L) in T2DM with CHD group were higher than those in T2DM without CHD((78.51±4.81)years old, (10.66±0.67)years, (6.84±2.19) mmol/L, (7.02±2.15)%, 63.03%(104/165), (0.21±0.04 ) mg/L, 13.33%(22/165), (83.01±14.40) μmol/L, (6.79±2.89) mmol/L, (333.56±95.15) μmol/L ), and the differences were statistically significant( t=15.908, t=2.042, t=3.055, t=3.088, χ 2=23.828, t=5.059, χ 2=42.098, t=2.401, t=4.188, t=4.075; all P<0.05). The total bilirubin(TBil)(8.80(6.60, 11.60) μmol/L), glomerular filtration rate(GFR)((76.49±29.80) mL/(min·1.75 m 2)) in T2DM with CHD group were lower than those in T2DM without CHD group (11.25(8.23, 15.28) μmol/L, (91.81±28.31) mL/(min·1.75 m 2)), the differences were statistically significant( Z=2.304, t=5.126; all P<0.001). The total cholesterol((3.84±0.85) mmol/L), low-density lipoprotein cholesterol(LDL-C)((2.12±0.68 ) mmol/L) in T2DM with CHD group were lower than those in T2DM without CHD group((4.10±1.00) mmol/L, (2.45±0.85) mmol/L), the differences were statistically significant( t=2.828, 4.156; all P<0.05). The rate of starting lipid-lowering and stable plaque treatment in T2DM with CHD group (82.57%(199/261))was higher than that in T2DM without CHD group(42.42%(70/165)), and the difference was statistically significant (χ 2=70.614, P<0.001). Influenced by lipid-lowering therapy, the total cholesterol and LDL-C in T2DM patients with CHD were significantly decreased.Logistic regression analysis showed that age elevated( OR 1.346, 95% CI 1.263-1.434, P<0.001), elevated hemoglobin a1c concentration( OR 1.427, 95% CI 1.140-1.785, P=0.002), complicated with hypertension( OR 3.534, 95% CI 1.684-7.418, P=0.001), elevated D-dimer concentration( OR 3.969, 95% CI 1.227-12.841, P=0.021)and elevated uric acid concentration( OR 1.005, 95% CI 1.001-1.008, P=0.006)were independent risk factors for CHD in elderly male patients with T2DM. Conclusion:Elderly patients with T2DM complicated with CHD are more likely to be in hypercoagulable state, more likely to have thromboembolic events, and more obvious renal function damage.Poor fasting blood glucose control and decreased total bilirubin concentration are the influencing factors of CHD in elderly male patients with T2DM.Age elevated, elevated hemoglobin a1c concentration, complicated with hypertension, elevated D-dimer concentration and elevated uric acid concentration are independent risk factors for CHD in elderly male patients with T2DM.

Objective To investigate the relation between the characteristics of CD8+T lymphocyte infiltration and the prognosis of triple-negative breast cancer patients. Methods We retrospectively analyzed the clinicopathological data of 126 patients with triple-negative breast cancer undergoing preoperative neoadjuvant chemotherapy. Immunohistochemical staining was used to analyze the relation between CD8+T lymphocyte infiltration and clinicopathological characteristics. Kaplan-Meier method was used to draw the survival curve, and Cox risk ratio regression model was used to analyze the prognostic factors affecting disease-free survival time (DFS). Results High-density CD8+Tils was associated with age < 60 years old, high pathological grade and high clinical stage (P < 0.05). The pCR rate of high-density CD8+Tils group was higher than that of the low-density group (66.7% vs. 19.8%, P=0.000). The median DFS of the high-density group was significantly longer than that of the low-density group (49 vs. 25 months, P < 0.05). Multivariate analysis showed that high pathological grade, tumor diameter > 2 cm, lymph node metastasis, vascular invasion and CD8+Tils low-density infiltration were factors for poor prognosis (P < 0.05), and CD8+Tils was an independent prognostic factor. Conclusion CD8+Tils may be an independent prognostic indicator for triple-negative breast cancer. The patients with high-density infiltration have high postoperative pCR rate, long DFS and better long-term efficacy.

Objective:To investigate the application of sentinel lymph node biopsy (SLNB) in early-staged cervical cancer by laparoscopy.Methods:It was a prospective, single-arm, single-center clinical study. Seventy-eight cases of cervical cancer patients were collected from July 2015 to December 2018 at the Fourth Hospital of Hebei Medical University. All the patients were injected with tracer into the disease-free block of cervical tissue after anesthesia by the same surgeon who learned sentinel lymph node (SLN) mapping technique in Memorial Sloan-Kettering Cancer Center, and underwent SLN mapping followed by complete pelvic lymphadenectomy. Moreover, all the dissected lymph nodes were stained with hematoxylin eosin staining (HE) pathological examination. Besides, the negative SLN on hematoxylin-eosin staining were detected by immunohistochemistry cytokeratin staining micro-metastasis. To analyze the distribution, detection rate, false negative rate the sensitivity and negative predictive value of the SLN in early-staged cervical cancer by laparoscopy, and explore the value of SLN mapping in predicting the lymph nodes metastasis in early-staged cervical cancer.Results:The overall detection rate of SLN in cervical cancer was 99% (77/78), bilateral detection rate was 87% (68/78). The average of 12.4 lymph node (LN) and 3.6 SLN were dissected for each patients each side. SLN of cervical cancer were mainly distributed in the obturator space (61.5%, 343/558), followed by external iliac (23.5%, 131/558), common iliac (7.3%, 41/558), para-uterine (3.8%, 21/558), internal iliac (2.2%, 12/558), para abdominal aorta (1.1%, 6/558), and anterior sacral lymphatic drainage area (0.7%, 4/558). Fourteen cases of LN metastasis were found among all 78 cases. There were a total of 38 positive LN, including 26 SLN metastasis and 12 none sentinel LN metastasis. Through immunohistochemical staining and pathological ultra-staging, 1 SLN was found to be isolated tumor cells (ITC), and 5 SLNs were found to be micro-metastases (MIC), accounting for 23% (6/26) of positive SLN. SLN mapping with pathological ultra-staging improved the prediction of LN metastasis in cervical cancer (2/14). Metastatic SLN mainly distributed in the obturator space (65%, 17/26), peri-uterine region (12%, 3/26), common iliac region (15%, 4/26), and external iliac region (8%, 2/26). The consistency of the diagnosis of lymph node metastasis by SLN biopsy and postoperative retroperitoneal lymph node metastasis showed that the Kappa value was 1.000 ( P<0.001), indicated that the metastasis status of SLN and retroperitoneal lymph node were completely consistent. The sensitivity, specificity, accuracy, false-negative rate, and negative predictive value of SLN biopsy in the diagnosis of lymph node metastasis were 100%, 100%, 100%, 0, and 100%, respectively. Conclusions:SLN in early-staged cervical cancer patients were mainly distributed in the obturator and external iliac space, pathalogical ultra-staging of SLN could improve the prediction of LN metastasis. Intraoperative SLN mapping is safe, feasible and could predict the state of retroperitoneal LN metastasis in early-staged cervical cancer. SLNB may replace systemic pelvic lymphadenectomy.

Microenvironmental biophysical factors play a fundamental role in controlling cell behaviors including cell morphology, proliferation, adhesion and differentiation, and even determining the cell fate. Cells are able to actively sense the surrounding mechanical microenvironment and change their cellular morphology to adapt to it. Although cell morphological changes have been considered to be the first and most important step in the interaction between cells and their mechanical microenvironment, their regulatory network is not completely clear. In the current study, we generated silicon-based elastomer polydimethylsiloxane (PDMS) substrates with stiff (15:1, PDMS elastomer vs. curing agent) and soft (45:1) stiffnesses, which showed the Young's moduli of ~450 kPa and 46 kPa, respectively, and elucidated a new path in cytoskeleton re-organization in chondrocytes in response to changed substrate stiffnesses by characterizing the axis shift from the secreted extracellular protein laminin β1, focal adhesion complex protein FAK to microfilament bundling. We first showed the cellular cytoskeleton changes in chondrocytes by characterizing the cell spreading area and cellular synapses. We then found the changes of secreted extracellular linkage protein, laminin β1, and focal adhesion complex protein, FAK, in chondrocytes in response to different substrate stiffnesses. These two proteins were shown to be directly interacted by Co-IP and colocalization. We next showed that impact of FAK on the cytoskeleton organization by showing the changes of microfilament bundles and found the potential intermediate regulators. Taking together, this modulation axis of laminin β1-FAK-microfilament could enlarge our understanding about the interdependence among mechanosensing, mechanotransduction, and cytoskeleton re-organization.
Cell Adhesion ; Chondrocytes ; Cytoskeleton/metabolism* ; Elastomers/metabolism* ; Laminin/metabolism* ; Mechanotransduction, Cellular

Objective: To understand the novel bat-borne hantavirus on Yunnan province. Methods: Eighty-four bats were collected from Puer, Yunnan from July to August in 2016. A hantavirus, named as DodeHV, was identified in Pomona Roundleaf Bat by high-throughput sequencing technology. Nested or Semi-nested reverse transcription-polymerase chain reaction was used to confirm and screen DodeHV, as well as to amplify its genome sequence by specific primers. Then homology and phylogenetic analysis of all three segments were conducted by using software MegAlign and MEGA 6.0. Results: All the positive individuals are Pomona Roundleaf bats by PCR in this study and 4 out of 67 (5.97%) Pomona Roundleaf bats were positive. The complete ORF of DodeHV S Segment, the most sequence of M and the complete sequence of L segment were obtained. It shared the highest homology with the strain XSV-VN1982B4 found on Phu Tho Province, Viet- Nam in 2013, the nucleotide sequence identities of S, M, L segment compared with XSV was 79.0%, 79.2%, 79.9% respectively, and its amino acid sequence identities was 93.4%, 94.8%, 96.6% respectively. Meanwhile, phylogenetic analysis showed that DodeHV was also closely related to strain XSV-VN1982B4. Conclusions The discovery of DodeHV enriched the virus reservoir of our country which is meaningful to public health. It is indicated that there is potential risk of bat-borne DodeHV at cross-border infection at frontier.

0bjective To identify the molecular structure and genome homology characteristics of a GV murine norovirus (MNV) from wild mice.Methods: Nucleic acids were extracted from collected mouse intestinal specimens and subjected to high-throughput sequencing. The whole genome sequence was obtained by RT-PCR and RACE method . Results: The MNV genome was 7, 832 bp in length, with 76.8%-78.1% nucleotide identity to other selected murine norovirus strains in GenBank. The P2 region, the hypervariable area in viral protein 1 gene shows highest heterology, and the identity was among 60.9%-78.1%, lower than other genes regions. The result demonstrated the higher variability of this newly isolated WNV strain. Conclusions The wild-type MNV isolated in this study is first reported in China, which enriched the domestic MNV gene pool and provided a reference for understanding the molecular genetic characteristics and evolutionary origin of MNV in China.

Objective: To investigate the clinical characteristics of secondary hemophagocytic lymphohistiocytosis (sHLH) complicated with capillary leak syndrome (CLS) . Methods: The clinical and laboratory data of 87 sHLH patients, who were treated in our hospital between January 2015 and December 2017, were retrospectively analyzed. Depending on whether they were complicated with CLS, 21 sHLH patients were classified as the CLS-sHLH group, while 66 were classified as the non-CLS-sHLH group. The differences of clinical manifestations, laboratory tests, treatment and prognosis between the two groups were compared. Results: There was no significant difference in the etiology of sHLH between the CLS-sHLH group and the non-CLS-sHLH group (P>0.05) . The neutrophil, fibrinogen and albumin levels in the CLS-sHLH group were lower than those in the non-CLS-sHLH group, while the triacylglycerol levels were higher than those in the non-CLS-sHLH group (P<0.05) . Varying degrees of edema, weight gain, hypotension, hypoproteinemia, oliguria and multiple serous effusions were observed in the CLS-sHLH group. Among them, there were 15 patients that CLS get improved, and the medial time of improvement was 7 (5-14) days. The other 6 patients did not get remission, while they died within 6-30 days. The median overall survival of the CLS-sHLH group was lower than that of the non-CLS-sHLH group (75 days vs not reached, P=0.031) . Conclusions There may be no correlation between the cause of sHLH and the occurrence of CLS. Severity of neutropenia, fibrinogen and albumin levels, and triglyceride levels may be accompanied for sHLH patients complicated with CLS. Patients with sHLH who complicated with CLS have a poor prognosis. Active treatment of HLH and its primary disease, reasonable fluid replacement and oxygen supply are crucial, which can effectively control disease progression.

Engineered probiotics can serve as therapeutics based on their ability of produce recombinant immune-stimulating properties. In this study, we built the recombinant Bacillus subtilis WB800 expressing antimicrobial peptide KR32 (WB800-KR32) using genetic engineering methods and investigated its protective effects of nuclear factor-E2-related factor 2 (Nrf2)‍-Kelch-like ECH-associated protein 1 (Keap1) pathway activation in intestinal oxidative disturbance induced by enterotoxigenic Escherichia coli (ETEC) K88 in weaned piglets. Twenty-eight weaned piglets were randomly distributed into four treatment groups with seven replicates fed with a basal diet. The feed of the control group (CON) was infused with normal sterilized saline; meanwhile, the ETEC, ETEC+WB800, and ETEC+WB800-KR32 groups were orally administered normal sterilized saline, 5×10¹⁰ CFU (CFU: colony forming units) WB800, and 5×10¹⁰ CFU WB800-KR32, respectively, on Days 1‍‒‍14 and all infused with ETEC K88 1×10¹⁰ CFU on Days 15‍‒‍17. The results showed that pretreatment with WB800-KR32 attenuated ETEC-induced intestinal disturbance, improved the mucosal activity of antioxidant enzyme (catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx)) and decreased the content of malondialdehyde (MDA). More importantly, WB800-KR32 downregulated genes involved in antioxidant defense (GPx and SOD1). Interestingly, WB800-KR32 upregulated the protein expression of Nrf2 and downregulated the protein expression of Keap1 in the ileum. WB800-KR32 markedly changed the richness estimators (Ace and Chao) of gut microbiota and increased the abundance of Eubacterium_rectale_ATCC_33656 in the feces. The results suggested that WB800-KR32 may alleviate ETEC-induced intestinal oxidative injury through the Nrf2-Keap1 pathway, providing a new perspective for WB800-KR32 as potential therapeutics to regulate intestinal oxidative disturbance in ETEC K88 infection.
Animals ; Swine ; Enterotoxigenic Escherichia coli ; Kelch-Like ECH-Associated Protein 1 ; Bacillus subtilis ; NF-E2-Related Factor 2 ; Antioxidants ; Oxidative Stress