Genetic variants in Mammalian STE20-like protein kinase 2 were associated with risk of NSCL/P.

AIM: Mammalian STE20-like protein kinase 2 (MST2) plays an important role in apoptosis and the development of many disorders. Here, we aim to explore if genetic variants in MST2 are associated with the risk of non-syndromic cleft lip with or without palate (NSCL/P). MATERIALS AND METHODS: The association study was performed in a two-stage study of 1,069 cases and 1,724 controls to evaluate the association between genetic variants in the MST2 and NSCL/P risk. The potential function of the candidate single nucleotide polymorphism (SNP) was predicted using HaploReg, RegulomeDB, and public craniofacial histone chromatin immunoprecipitation sequencing (ChIP-seq) data. Haploview was used to perform the haplotype of risk alleles. The expression quantitative trait loci (eQTL) effect was assessed using the Genotype-Tissue Expression (GTEx) project. Gene expression in mouse embryo tissue was performed using data downloaded from GSE67985. The potential role of candidate gene in the development of NSCL/P was assessed by correlation and enrichment analysis. RESULTS: Among SNPs in MST2, rs2922070 C allele (Pmeta = 2.93E-04) and rs6988087 T allele (Pmeta = 1.57E-03) were linked with significantly increased risk of NSCL/P. Rs2922070, rs6988087 and their high linkage disequilibrium (LD) SNPs constituted a risk haplotype of NSCL/P. Individuals carrying 3-4 risk alleles had an elevated risk of NSCL/P compared to those who carried less risk alleles (P = 2.00E-04). The eQTL analysis revealed a significant association between these two variants and MST2 in muscle tissue of the body. The MST2 expressed during mouse craniofacial development and over-expressed in the human orbicularis oris muscle (OOM) of NSCL/P patients compared to controls. MST2 was involved in the development of NSCL/P by regulating the mRNA surveillance pathway, the MAPK signaling pathway, the neurotrophin signaling pathway, the FoxO signaling pathway and the VEGF signaling pathway. CONCLUSION: MST2 was associated with the development of NSCL/P.

Integrating GWAS and eQTL to predict genes and pathways for non-syndromic cleft lip with or without palate.

OBJECTIVE: To explore susceptibility genes and pathways for non-syndromic cleft lip with or without cleft palate (NSCL/P). MATERIALS AND METHODS: Two genome-wide association studies (GWAS) datasets, including 858 NSCL/P cases and 1,248 controls, were integrated with expression quantitative trait loci (eQTL) dataset identified by Genotype-Tissue Expression (GTEx) project in whole-blood samples. The expression of the candidate genes in mouse orofacial development was inquired from FaceBase. Protein-protein interaction (PPI) network was visualized to identify protein functions. Go and KEGG pathway analyses were performed to explore the underlying risk pathways. RESULTS: A total of 233 eQTL single-nucleotide polymorphisms (SNPs) in 432 candidate genes were identified to be associated with the risk of NSCL/P. One hundred and eighty-three susceptible genes were expressed in mouse orofacial development according to FaceBase. PPI network analysis highlighted that these genes involved in ubiquitin-mediated proteolysis (KCTD7, ASB1, UBOX5, ANAPC4) and DNA synthesis (XRCC3, RFC3, KAT5, RHNO1) were associated with the risk of NSCL/P. GO and KEGG pathway analyses revealed that the fatty acid metabolism pathway (ACADL, HSD17B12, ACSL5, PPT1, MCAT) played an important role in the development of NSCL/P. CONCLUSIONS: Our results identified novel susceptibility genes and pathways associated with the development of NSCL/P.

Liquid Chromatography-Mass Spectrometry-Based Plasma Metabolomics Study of the Effects of Moxibustion with Seed-Sized Moxa Cone on Hyperlipidemia.

Hyperlipidemia (HLP) is a disorder with disturbed lipid metabolism. HLP is a major risk factor in cardiovascular diseases, atherosclerosis, diabetes mellitus, and coronary heart disease. This study focuses on understanding the effects of moxibustion with a seed-sized moxa cone on HLP and the potential metabolic pathways associated with HLP. An automatic analyzer was used to measure the levels of total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) in healthy controls (HCs), HLP patients, and in patients before moxibustion with seed-sized moxa cone treatment (BMT) and after moxibustion treatment (AMT). Liquid chromatography-mass spectrometry and pathway analyses were performed using differential plasma metabolites derived from the HC, HLP, BMT, and AMT groups. Higher levels of TC, TG, and LDL-C and lower levels of HDL-C were detected in HLP patients than in HCs. The levels of TC and TG were significantly decreased in the AMT group compared to those of the BMT group. A total of 87 differential metabolites were identified from the HLP vs HC samples and 51 for the AMT vs BMT samples. Of these, 21 terms were shared. The differential metabolites in both the HLP vs HC and AMT vs BMT groups were significantly enriched in the glycerophospholipid and sphingolipid metabolism pathways. We suggest that moxibustion with seed-sized moxa cone treatment is effective against hyperlipidemia by altering the levels of TC and TG, which might be regulated by glycerophospholipid and sphingolipid metabolism.

γδ T cells in hepatocellular carcinoma patients present cytotoxic activity but are reduced in potency due to IL-2 and IL-21 pathways.

Hepatocellular carcinoma (HCC) is the most common primary liver carcinoma and has one of the highest mortality rates of all cancers. The γδ T cells could infiltrate HCC and have demonstrated potent tumor-killing capacity. Here, we found that in peripheral blood, the vast majority of γδ T cells were Vδ2 T cells. In HCC patients, the frequency of Vδ2 T cells was significantly lower than in controls. γδ T cells that were harvested directly ex vivo possessed very limited capacity to eliminate Zol-loaded HCC cell lines, even at a high effector to target ratio. In vitro expansion with Zol could significantly increase the capacity of γδ T cells to eliminate HCC cell lines. But even with in vitro expansion, the γδ T cells from HCC patients presented significantly lower cytotoxic capacity than the γδ T cells from healthy individuals. The expression of IL-2 and IL-21 by γδ T cells was significantly lower in HCC patients than in control volunteers. Supplementing recombinant human IL-2 and IL-21 in the in vitro expansion culture increased the cytotoxic capacity of γδ T cells. In addition, the frequency of PD-1+ γδ T cells was significantly higher in HCC patients than in controls ex vivo, and was significantly elevated after in vitro expansion. Hep3B and HepG2 did not express PD-L1, while a small fraction of SNU-398 expressed PD-L1. Interestingly, co-incubation with γδ T cell elevated PD-L1 expression in HCC cell lines. Blocking PD-1 during in vitro expansion stage significantly elevated cytotoxicity toward all the HCC cell lines, while blocking PD-1 during the cytotoxicity assay significantly elevated cytotoxicity toward HepG2 and SNU-398, but not toward Hep3B. Overall, these results demonstrated that the circulating γδ T cells in HCC patients were reduced in cytotoxic capacity, possibly associated with the lack of IL-2 and IL-21 production and PD-1 upregulation.

Epalrestat protects against diabetic peripheral neuropathy by alleviating oxidative stress and inhibiting polyol pathway.

Epalrestat is a noncompetitive and reversible aldose reductase inhibitor used for the treatment of diabetic neuropathy. This study assumed that epalrestat had a protective effect on diabetic peripheral nerve injury by suppressing the expression of aldose reductase in peripheral nerves of diabetes mellitus rats. The high-fat and high-carbohydrate model rats were established by intraperitoneal injection of streptozotocin. Peripheral neuropathy occurred in these rats after sustaining high blood glucose for 8 weeks. At 12 weeks after streptozotocin injection, rats were intragastrically administered epalrestat 100 mg/kg daily for 6 weeks. Transmission electron microscope revealed that the injuries to myelinated nerve fibers, non-myelinated nerve fibers and Schwann cells of rat sciatic nerves had reduced compared to rats without epalrestat administuation. Western blot assay and immunohistochemical results demonstrated that after intervention with epalrestat, the activities of antioxidant enzymes such as superoxide dismutase, catalase and glutathione peroxidase gradually increased, but aldose reductase protein expression gradually diminished. Results confirmed that epalrestat could protect against diabetic peripheral neuropathy by relieving oxidative stress and suppressing the polyol pathway.

Interactions Between Targeted Ultrasound Contrast Agents With Anti-Human Interleukin 8 Monoclonal Antibody and Activated Endothelial Cells.

The study aimed to explore the role of interleukin 8 (IL-8) in atherosclerotic plaques and develop a new method for the evaluation of endothelial function by assessing the interactions between the injured endothelial cells and the targeted ultrasound agent that carried anti-human IL-8 monoclonal antibody. Anti-human IL-8 monoclonal antibodies were associated to the shells of SonoVue microbubbles by covalent conjugation technology. The specific interaction between the microbubbles and the normal or injured endothelial cells was observed using an inverted microscope. The microbubble adherence was quantified by calculating the ratio of the adherent microbubbles to endothelial cells. The results were compared with the control microbubbles. There were rare adherences of control microbubbles to the normal or injured endothelial cells, whereas the targeted microbubbles could adhere to endothelial cells well. Importantly, compared with the normal endothelial cells, a significantly higher number of targeted microbubbles bound to the injured endothelial cells. The ultrasound agents with anti-human IL-8 monoclonal antibody can specifically bind to the injured endothelial cell, which provides a new insight to the noninvasive detection of endothelial dysfunction using ultrasound imaging techniques.

Transplantation of umbilical cord mesenchymal stem cells via different routes in rats with acute liver failure.

OBJECTIVE: This study aimed to compare the therapeutic efficacy of transplantation of human umbilical cord mesenchymal stem cells (hUCMSC) in different routes in acute hepatic failure (ALF) in rats. METHODS: hUCMSCs were isolated and identified by detection of surface antigens via flow cytometry. In T group and H group, ALF rats received hUCMSC transplantation through the tail vein and intrahepatic injection, respectively. In hUCMSC group, healthy rats received hUCMSCs transplantation via the tail vein. In ALF group, rats received injection of normal saline through the tail vein. RESULTS: The TBil and ALT in ALF rats with and without transplantation were significantly higher than in healthy rats (P<0.05). HE staining of the liver showed obvious hepatocyte regeneration and reduced infiltration of inflammatory cells, and liver pathology was improved in T group and H group as compared to ALF group. At 3 d after transplantation, CK18 expression was detectable in both H group and T group. At 1 w and 2 w, the mRNA expressions of CK8, CK18 and AFP in H group and T group were significantly different from those in ALF group (P<0.05). The liver function and differentiation of stem cells were comparable between H group and T group (P>0.05). CONCLUSION: hUCMSCs transplantation can improve the liver function and promote the liver repair following ALF. hUCMSCs transplantation via tail vein has similar therapeutic efficacy to that through intrahepatic injection.

Effect of bone marrow mesenchymal stem cells transplantation on the serum and liver HMGB1 expression in rats with acute liver failure.

OBJECTIVE: This study aimed to investigate the effect of bone marrow mesenchymal stem cells (BMSCs) transplantation on the expression of high mobility group box 1 protein (HMGB1) in the serum and liver of rats with acute liver failure (ALF). METHODS: Healthy male SD rats were randomly divided into control group, ALF group and BMSCs group. ALF was induced by intraperitoneal injection of 900 mg/kg D-GalN and 10 µg/kg LPS. In BMSCs group, rats received BMSCs (1.0×10(7)) transplantation via the tail vein at 2 h after ALF induction. RESULTS: Intraperitoneal injection of 900 mg/kg D-GalN and 10 µg/kg LPS was able to induce ALF in rats. In ALF group, serum ALT and AST increased gradually over time. At 72 h, the serum ALT and AST in BMSCs group were significantly different from those in ALF group. HMGB1 expression in the serum and liver remained at a low level at any time point in control group, but increased significantly in ALF group and BMSCs group. The serum and liver HMGB1 expression increased progressively in ALF group, but reduced gradually in BMSCs group. Significant difference in serum and liver HMGB1 expression was observed between ALF group and BMSCs group at 24 h and 72 h. In addition, there was marked difference in the survival rate among three groups at 24 h (χ (2) =21.098, P<0.01). CONCLUSION: BMSCs transplantation is able to improve the liver function and liver pathology in ALF rats and decrease the serum and liver HMGB1.

Effects of lip repair on maxillofacial morphology in patients with unilateral cleft lip with or without cleft palate.

Objective : To evaluate the effects of lip repair on maxillofacial development of patients with unilateral cleft lip with or without cleft palate. Design : Retrospective. Patients : A total of 75 patients were recruited, including 38 surgical patients with complete unilateral cleft lip and alveolus and 37 patients with complete unilateral cleft lip and palate who had lip but not palate repair. As controls, 38 patients with no cleft were selected. All subjects were divided according to two growth stages: before the pubertal peak (GS1) and after the pubertal peak (GS2). Interventions : Lateral cephalograms of all subjects were obtained. Main Outcome Measures : Cephalograms were analyzed and compared in the study and control groups. Results : The patients with unilateral cleft lip and palate in both GS1 and GS2 demonstrated an almost normal maxillary and mandibular growth with retroclined maxillary incisors. The patients with unilateral cleft lip and palate showed a shorter length of maxilla, a more clockwise-rotated mandible, and retroclined maxillary incisors. Conclusions : There was an almost normal maxillary and mandibular growth but retroclined maxillary incisors in patients with cleft lip with or without cleft palate who had received lip repair only, indicating that lip repair may not have a negative impact on the maxillofacial development and influences only the inclination of the maxillary incisors. The shorter anterior-posterior maxillary length and larger gonial angle in patients with unilateral cleft lip and palate compared with those in patients with unilateral cleft lip and alveolus suggest that these variations in maxillary and mandibular growth may be a consequence of the cleft itself.

Biomechanical effects on maxillary protraction of the craniofacial skeleton with cleft lip and palate after alveolar bone graft.

OBJECTIVE: To explore the biomechanical effects of maxillary protraction on the craniofacial skeleton in patients with unilateral cleft lip and palate (UCLP) after alveolar bone graft (ABG) and resorption of ABG, thus to provide theoretical basis for the clinical application of maxillary protraction, which can improve the facial deformity of the UCLP patients. METHODS: A finite element model of a UCLP patient's skull was generated using data from spiral computed tomographic (CT) scans. Based on this finite element model, another 6 ABG finite element models were constructed to simulate ABG and resorption of ABG, respectively (nonresorption model, upper one-third resorption of the grafted bone model, upper two-thirds resorption of the grafted bone model, lower one-third resorption of the grafted bone model, lower two-thirds resorption of the grafted bone model, upper one-third and lower one-third resorption of the grafted bone model). Two additional models were developed to simulate maxillary protraction with expansion and maxillary protraction alone. All models were loaded with orthopedic force (30 degrees downward and forward to the occlusal plane, 500 g per side) on the region of alveolar of maxillary canine. RESULTS: Before ABG, the cleft side showed larger displacement than the noncleft side, when it came to the stress distribution in the craniofacial suture, it showed an asymmetric pattern as well. After ABG, the displacement difference between the cleft side and the noncleft side decreased, and the stress distribution in the craniofacial suture showed more symmetric than that before ABG. The pterygopalatine suture obtained the largest value, followed by zygomaticotemporal, zygomaticomaxillary, and zygomaticofrontal sutures among the observed sutures. Higher stresses and pronounced forward displacement were generated in the craniofacial sutures after maxillary protraction with expansion. CONCLUSIONS: Maxillary protraction after ABG performed a more favorable outcome. Among the ABG models, nonresorption model showed the best effect after loading maxillary protraction force, and resorption in the lower region of the grafted bone showed a better effect than resorption in the upper region of the grafted bone. Maxillary expansion could effectively facilitate the orthopedic of the maxillary protraction presumably.

Assessment of atherosclerotic plaques in the rabbit abdominal aorta with interleukin-8 monoclonal antibody-targeted ultrasound microbubbles.

In this study, we aimed to prepare a neovascularization-relevant inflammatory cytokine-targeted ultrasound contrast agent and apply it in the ultrasound imaging of atherosclerotic plaque. An interleukin-8 (IL-8) monoclonal antibody was conjugated to SonoVue microbubbles using the N-succinimidyl-3-(2-pyridyldithio)propionate cross-linking method. Then, a prepared IL-8-targeted contrast agent was used for contrast-enhanced ultrasound (CEU) to detect rabbit abdominal aorta atherosclerotic plaque and to investigate the imaging characteristics of atherosclerotic plaque with the contrast agent. We found that an IL-8 monoclonal antibody can be successfully coupled to SonoVue microbubbles with stable biological characteristics. CEU with this IL-8-targeted contrast agent can increase the atherosclerotic plaque detection sensitivity, with stronger echo, so that three more plaques were detected compared with using non-targeted SonoVue microbubbles. Thus, an inflammatory cytokine-targeting ultrasound contrast agent carrying IL-8 monoclonal antibody can provide unique advantages for researching the characteristics of atherosclerotic plaque.

Electroacupuncture regulates glucose-inhibited neurons in treatment of simple obesity.

The glucose-inhibited neurons present in the lateral hypothalamic area are regarded as glucose detectors. This structure is involved in the regulation of food intake through extracellular blood glucose concentrations, and plays a crucial role in obesity onset. In the present study, obesity models established with high fat feeding were treated with electroacupuncture at Zusanli (ST36)/Inner Court (ST44) on the left side and Tianshu (ST25) bilaterally. We found that electroacupuncture could effectively reduce body weight and the fat-weight ratio, and decrease serum leptin, resistin, tumor necrosis factor alpha, and neuropeptide Y levels, while increase serum adiponectin and cholecystokinin-8 levels. This treatment altered the electrical activity of glucose-inhibited neurons in the lateral hypothalamic area, with electroacupuncture at Zusanli/Inner Court exerting an inhibitory effect, while electroacupuncture at bilateral Tianshu exerting an excitatory effect. These data suggest that electroacupuncture at the lower limbs and abdominal cavity is an effective means for regulating the activity of glucose-inhibited neurons in the lateral hypothalamic area and for improving the secretory function of adipose tissue.

[125I seed implantation in sphincter preservation for treatment of low rectal cancer].

OBJECTIVE: To evaluate the effects of (125)I seed implantation in sphincter preservation for treatment of low rectal cancer. METHODS: Seventy-six patients with low rectal cancer were randomly divided into 2 group: group A, 17 males and 13 females, aged 48.5 +/- 2.4, receiving rectostomy and anal sphincter preservation and group B, 24 males and 22 females, aged 49.4 +/- 2.6, receiving modified TME and anal sphincter preservation combined with brachytherapy by (125)I seed implantation. Two to four weeks after operation chemotherapy with 5-FU/CF were performed. Follow-up was carried out 6, 12, 24, and 36 months after operation. RESULTS: The local recurrence rates 6, 12, 24, and 36 months after operation were 0%, 11.1%, 14.3%, and 23% respectively in the group A, and all 0% in the group B (P < 0.05 for the rate 36 months later). The survival rates 6, 12, 24, and 36 months after operation were 100%, 100%, 85.7%, and 76.7% respectively in the group A, and were 100%, 100%, 97.1%, and 93% respectively in the group B (P < 0.05 for the rate 36 months later). The functions of defecation and erection were better in the group B and the symptom of pain was improved better in the group A too (all P < 0.05). CONCLUSIONS: Safe, simple, and effective, surgery with sphincter preservation combined with brachytherapy in low rectal cancer is one of the ideal methods for treatment of low rectal cancer.

[Iodine-125 interstitial brachytherapy for malignant tumor].

OBJECTIVE: To assess the clinical effects of (125)I interstitial brachytherapy for malignant tumors. METHODS: One hundred and twelve patients with malignant tumors of stage II stage and III under went radical resection combined with (125)I intraoperative implantation. Seven days and three month after operation WBC count and immune markers were observed. Blood biochemistry ultrasonography and X-ray were performed to observe recurrence and metastasis of tumors per three months. RESULTS: In the 112 patients, 3 died of tumor recurrence in six months, and others survived with the longest time of 35 months. CONCLUSION: (125)I interstitial brachytherapy for malignant tumor is simple, safe and effective.