Regulatory T cells differentiation in visceral adipose tissues contributes to insulin resistance by regulating JAZF-1/PPAR-γ pathway.

Regulatory T cell (Treg) activity and differentiation in visceral adipose tissue (VAT) play an important role in inhibiting chronic inflammation and insulin resistance. Whether JAZF-1 and PPAR-γ mediate VAT Treg differentiation to promote the inhibition of chronic inflammation and insulin resistance remains unclear. Here, we investigated the roles of JAZF-1 and PPAR-γ in VAT Treg differentiation, inflammation and insulin resistance using a transgenic mouse model. First, we determined that the levels of glucose and insulin biochemical markers in the JAZF-1 transgenic general feeding or high-fat groups were lower than those in the wild-type general feeding or high-fat groups. Second, the levels of CD4+ , CD25+ , and FOXP3+ differentiation markers in the JAZF-1 transgenic general feeding or high-fat groups were significantly higher than those in the wild-type groups. PPAR-γ inhibition was associated with low levels of CD4+ , CD25+ and FOXP3+ differentiation markers. Third, the levels of TNF-α, IL-1ß and IL-6 in the JAZF-1 transgenic groups were lower than those in the wild-type groups, whereas IL-10 and TGF-ß levels were higher in the JAZF-1 transgenic groups than in the wild-type groups. After using the PPAR-γ inhibitor, we observed that TNF-α, IL-1ß and IL-6 increased, while IL-10 and TGF-ß decreased. We found that JAZF-1 and PPAR-γ could promote Tregs differentiation and regulate insulin resistance by synergistically decreasing the expression levels of TNF-α, IL-1ß and IL-6 and increasing those of IL-10 and TGF-ß.

Eupalinolide A induces autophagy via the ROS/ERK signaling pathway in hepatocellular carcinoma cells in vitro and in vivo.

Hepatocellular carcinoma is the most common primary malignancy of the liver. The current systemic drugs used to treat hepatocellular carcinoma result in low overall survival time. It has therefore been suggested that new small­molecule drugs should be developed for treating hepatocellular carcinoma. Eupatorium lindleyanum DC. (EL) has been used to treat numerous diseases, particularly respiratory diseases; however, to the best of our knowledge, studies have not yet fully elucidated the effect of EL on hepatocellular carcinoma. In the present study, the effect of eupalinolide A (EA), one of the extracts of EL, was evaluated on tumor growth in a xenograft model of human hepatocellular carcinoma cells, and on the proliferation and migration of hepatocellular carcinoma cell lines. Cell cycle progression and the type of cell death were then evaluated using the Cell Counting Kit 8 assay, flow cytometry, electron microscopy and western blotting. EA significantly inhibited cell proliferation and migration by arresting the cell cycle at the G1 phase and inducing autophagy in hepatocellular carcinoma cells. EA­induced autophagy was mediated by reactive oxygen species (ROS) and ERK signaling activation. Specific inhibitors of ROS, autophagy and ERK inhibited EA­induced cell death and migration. In conclusion, the present study revealed that EA may inhibit the proliferation and migration of hepatocellular carcinoma cells, highlighting its potential as a promising antitumor compound for treating hepatocellular carcinoma.

Effects of Adenovirus-Mediated Overexpression of JAZF1 on Chronic Inflammation: An In Vitro and In Vivo Study.

BACKGROUND Insulin sensitivity and inflammation can be affected by juxtaposition with another zinc finger gene 1 (JAZF1), but its precise role in chronic inflammation is unclear. In this study, JAZF1-overexpression adenovirus plasmids were transfected into macrophages, CD4⁺ T cells, and C57BL/6J mice to assess the role of JAZF1 in chronic inflammation. MATERIAL AND METHODS JAZF1 was cloned into an adenovirus skeleton plasmid and transfected in HEK293 cells to package and enrich the virus particles. In vitro, the JAZF1 overexpression adenovirus vector (PAD-JAZF1) was cultured with peritoneal macrophages and peripheral blood CD4⁺ T cells of C57BL/6J mice, and samples were evaluated using flow cytometry. In vivo, PAD-JAZF1 was introduced into C57BL/6J mice, and livers were collected to evaluate factors related to inflammation by hematoxylin & eosin and immunohistochemical staining. RESULTS In vitro, PAD-JAZF1 decreased total macrophages, CD11c⁺ macrophages, and the secretion of proinflammatory cytokines, but increased CD206⁺ macrophages. It also decreased total CD4⁺T cells, active T cells, memory T cells, and the secretion of IL-6, IL-10, and IFN-γ, but increased Treg cells and restrictive T cells. In vivo, compared to those in the control group transfected with the adenovirus skeleton vector, mice transfected with the PAD-JAZF1 recombinant adenovirus had fewer CD11c⁺ ATMs and CD4⁺ T cells, lower levels of TNF-alpha and IL-6, and higher IL-10 concentrations in the liver. CONCLUSIONS These findings indicate that JAZF1 limits chronic inflammation by reducing macrophage and CD4⁺T cell populations, altering subtype differentiation, and regulating the secretion of immune-related factors.

[Identification of the interactions between JTV1 and NLS-RAR alpha in vivo and in vitro].

OBJECTIVE: To identify the interactions between JTV1 and NLS-RAR alpha by Yeast two-hybrid and co-immunoprecipitation. METHODS: The plasmids of bait-protein and JTV1 protein were cotransformed into yeast AH109 to investigate their interactions in vivo. Tagged fusion protein eukaryotic expression vectors were constructed and then cotransfected into human embryo kidney 293 cells. Co-immunoprecipitation was used to investigate the interactions between NLS-RAR alpha and JTV1 in vitro. RESULTS: Blue clones were found in QDO/X-alpha-gal plates. Eukaryotic expression vectors were co-transfected into HEK 293 cells. The HA-NLS-RAR alpha protein was immunoprecipitated by anti-HA polyclonal antibody. Myc-JTV1 protein was detected by western blotting with anti-Myc monoclonal antibody from the immunoprecipitared complex. CONCLUSION: The interactions between NLS-RAR alpha and JTV1 are identified by Yeast two-hybrid and co-immunoprecipitation.

Use of a topical anesthetic cream (EMLA) to reduce pain after hemorrhoidectomy.

BACKGROUND AND OBJECTIVES: Hemorrhoidectomy usually leads to severe postoperative pain that often causes urinary retention. Topical EMLA cream (lidocaine 2.5% and prilocaine 2.5%) has been used extensively in the clinical setting. This prospective study tested the effectiveness of EMLA cream for postoperative pain control after hemorrhoidectomy. METHODS: Thirty patients admitted for hemorrhoidectomy were enrolled and randomly assigned into either a control group (n = 15) or EMLA group (n = 15). Postoperatively, the control group received approximately 5 g of neomycin ointment, and the EMLA group received approximately 5 g of EMLA. A visual analog scale (VAS) score was recorded on arrival in the postanesthesia recovery unit (PAR), after 2 hours in the PAR, on the first postoperative evening, and on the first postoperative morning. The requested frequency and dosage of meperidine, the first spontaneous voiding time, the frequency of single urinary catheterization, and a patient satisfaction score were also obtained. RESULTS: The VAS score and frequency and dosage of meperidine injections were significantly lower in the EMLA group than in the control group (P < .01). The voiding time was significantly later in the control group (P = .04). The frequency of single catheterization was significantly lower in the EMLA group than in the control group (P = .03). Patient satisfaction with postoperative pain control was significantly higher in the EMLA group than in the control group (P < .01). No systemic complications were observed. CONCLUSIONS: Topical EMLA cream decreased pain intensity and meperidine requests, reduced the frequency of single catheterizations, and improved patient satisfaction with postoperative pain management after hemorrhoidectomy in adults.

JAZF1 Inhibits Adipose Tissue Macrophages and Adipose Tissue Inflammation in Diet-Induced Diabetic Mice.

BACKGROUND: Juxtaposed with another zinc finger gene 1 (JAZF1) affects gluconeogenesis, insulin sensitivity, lipid metabolism, and inflammation, but its exact role in chronic inflammation remains unclear. This study aimed to examine JAZF1 overexpression in vivo on adipose tissue macrophages (ATMs). METHODS: Mouse models of high-fat diet- (HFD-) induced insulin resistance were induced using C57BL/6J and JAZF1-overexpressing (JAZF1-OX) mice. The mice were randomized (8-10/group) to C57BL/6J mice fed regular diet (RD) (NC group), C57BL/6J mice fed HFD (HF group), JAZF1-OX mice fed RD (NJ group), and JAZF1-OX mice fed HFD (HJ group). Adipose tissue was harvested 12 weeks later. ATMs were evaluated by flow cytometry. Inflammatory markers were evaluated by ELISA. RESULTS: JAZF1-OX mice had lower blood lipids, blood glucose, body weight, fat weight, and inflammatory markers compared with HF mice (all P < 0.05). JAZF1 overexpression decreased ATM number and secretion of proinflammatory cytokines. JAZF1 overexpression decreased total CD4+ T cells, active T cells, and memory T cells and increased Treg cells. JAZF1 overexpression downregulated IFN-γ and IL-17 levels and upregulated IL-4 levels. JAZF1 overexpression decreased MHCII, CD40, and CD86 in total ATM, CD11c+ ATM, and CD206+ ATM. CONCLUSIONS: JAZF1 limits adipose tissue inflammation by limiting macrophage populations and restricting their antigen presentation function.

Paravertebral transcutaneous electrical nerve stimulation reduces movement during general anesthesia with isoflurane.

We evaluated paravertebral transcutaneous electrical nerve stimulation (TENS) as a means of enhancing anesthesia during hysterectomy. Patients were randomly assigned to experimental (n = 21) and control (n = 20) groups. Anesthesia with isoflurane was performed uniformly for all patients. Paravertebral (T6 and T7) TENS (50 mA, 15 Hz, continuously) was applied in the experimental group. After 15 min of isoflurane, a lower abdominal, skin-to-adipose-tissue incision was made. Seventeen of 21 patients in the experimental group showed no arm or leg movements during the incision, compared to 8 with 20 patients in the control group (P = 0.007). TENS deserves further exploration as an adjunct technique for general anesthesia.

[Screening of target proteins interacting with dexamethasone derivates in vivo].

BACKGROUND AND OBJECTIVE: The anti-tumor activity of dexamethasone derivatives (9-fluoro-16alpha-methyl-11,17-dihydroxy-3-oxo-1,4-androsladiene-17beta-carboxylic acid) is superior to that of dexamethasone. This study was to screen the proteins interacting with dexamethasone derivates, thus to explore the anti-tumor mechanism of dexamethasone derivates in vivo. METHODS: The bait plasmid pGBKT7-GRalpha-LBD was constructed. Screening of the target proteins interacting with dexamethasone derivatives was performed by yeast three-hybrid technique using human K562 cell cDNA library. RESULTS: The bait plasmid was successfully constructed. It produced a 31 ku bait protein with no toxicity, leakage and self-activation. Thirty-seven positive clones which interacted with dexamethasone derivatives were obtained from human K562 cell cDNA library, 20 of which were identified by re-transforming into yeast AH109 cells. CONCLUSION: Twenty positive clones interacting with dexamethasone derivates are identified in vivo.

[Screening and identification of proteins interacting with RAR alpha-V via yeast two-hybrid system].

OBJECTIVE: To screen the protein interacting with retinoic acid receptor variant protein (RAR alpha-V) via the yeast two-hybrid technique (YTHT) and to find out the targets protein and study its biological function. METHODS: The bait vector of pGBKT7-RAR alpha-V was constructed for screening proteins interacting with RAR alpha-V in K562 cell cDNA expression library via YTHT. The protein-protein interaction was confirmed with re-transformation in yeast and GST pull-down in vitro. RESULTS: The bait vector was successfully constructed without toxicity, leakage and self-activation. Sixteen proteins were screened by YTHT and eight positive clones were identified by re-transformation in yeast. The interaction between RAR alpha-V and JTV-1 was confirmed by GST pull-down in vitro. CONCLUSIONS: There are some kinds of proteins interacting with RAR alpha-V in cell. The biological dysfunction caused by certain protein-protein interaction may be involved in the pathogenesis of leukemia.

A two-stage approach to positioning and identification of tracheal intubation using LED-based lightwand and acoustic models.

This paper proposes a two-stage approach to positioning and identifying tracheal intubations by computer-assisted diagnosis system. First, an innovated LED-based lightwand is developed for positioning the opening of the trachea and inserting the endotracheal tube rapidly. The proposed LED-based lightwand instrument significantly reduces the effects of temperature reaction without changing transillumination through the tissue. After intubation, breath sound analysis instrumentation is developed and assists for identifying the accurate tube position. To overcome the fast and overstepped variations of amplitude, a high sensitivity omni dimensional microphone and an automatic gain control device with linear phase property are calibrated and conducted into experiments of operating intubations. User-friendly interface software is also developed to analyze and transcribe the physiological characteristics of the collected breath sound corpus. Several subjective and objective experiments are performed to examine the practicability of the proposed approach and systems. The preliminary results show that the proposed LED-based lightwand instrument outperformed in the tip temperature without the influence on transillumination. The proposed parameters of average vibration parameter variance (AVPV) and frequency-domain energy variance (FEV) revealed the potential for distinguishing between the tracheal and esophageal intubations.

Earlier cessation of desflurane supply in closed-circuit anesthesia reduces emergence time in patients undergoing breast surgery.

BACKGROUND: Minimizing the time of anesthesia emergence can facilitate faster patient turnover in the operating rooms of a busy surgery center. According to Lin's new concept of inhalation uptake, after turning off the vaporizer under close-circuit anesthesia (CCA) with a very low fresh gas flow rate, the concentration of desflurane decreases at a slow rate. The aim of this study was to determine if earlier cessation of desflurane supply would shorten the emergence time and at the same time register the changes of desflurane concentration in the circuit after turning off the vaporizer. METHODS: 30 patients were randomly assigned to two groups, i.e., the control group and the study group. In the control group, the desflurane supply was continued up till the end of the operation, while in the study group the desflurane supply was cut off prior to the suturing the skin. In the study group, data regarding the hemodynamic changes, time from turning off desflurane with high flow washout to wakefulness, and the inspired as well as the expired desflurane concentrations at the low-flow anesthetic phase were collected. The time required from high flow washout to emergence was recorded in all patients. Inter-group and intra-group data were analyzed with nonparametric 2-independent-samples Mann-Whitney test and 2 related-samples Wilcoxon signed ranks test, respectively. RESULTS: Under CCA with similar surgical duration, the patients in the study group emerged from anesthesia significantly faster than those in the control group (5.6 +/- 1.9 min versus 8.8 +/- 2.3 min; P < 0.05), without molestation of stable hemodynamic signs. At the low-flow wash-in stage, the inspired desflurane concentrations were significantly higher than the expired ones from 0 to 2nd min; no significant difference was noted from the 3rd to 6th min, but after which the expired concentrations were significantly higher. Desflurane concentrations decreased most noticeably during the first 5 min (0.35 +/- 0.14%), and then the decrease was moderating from 6th to 10th min (0.21 +/- 0.58%) and staggered from 11th to 15th (0.14 +/- 0.06%). The mean duration of low flow wash was 25.6 +/- 11.6 min. No patient reported awareness during surgery. CONCLUSIONS: Ceasing desflurane supply earlier in CCA (250 mL/min) significantly shortens emergence time without significant hemodynamic changes.

Minimal dosage of tetracaine supplemented with epinephrine for spinal anesthesia in anorectal surgery.

BACKGROUND: Spinal anesthesia has been widely used in clinical setting with relatively high incidences of hypotension and bradycardia. Lowering the dosage of local anesthetics is one of the methods to mitigate the side effects. This study was to evaluate the feasibility of lowering the dosage of tetracaine in spinal anesthesia for patients undergoing anorectal surgery. METHODS: Thirty patients scheduled for anorectal surgery were studied. Patients were randomly divided into experiment (n = 15, 3.0 mg of tetracaine) and control groups (n = 15, 6.0 mg of tetracaine). The extent of analgesia was assessed by loss-of-sensation to pinprick. Dermatomic level of the sensory block was evaluated and recorded every minute for 10 minutes. BP and HR were recorded at 3-min interval for the 10-min in the study period. Numeric data were statistically analyzed with Student's t-test. The categorical data were compared using the chi-square test. P-value less than 0.05 was considered statistically significant. RESULTS: Ten min after the injection, the mean peak level of sensory block reached T12 for experiment and T9 for control groups. A noticeable difference in frequency of hypotension between two groups was found though it was not statistically significant (P = 0.08). Incidences of moderate bradycardia and severe bradycardia were similar in both groups, being 13.3% and 6.7% respectively. CONCLUSIONS: This study confirmed that lowering the dosage of tetracaine to 3.0 mg could equally provide adequate spinal anesthesia for anorectal surgery. The reduced dosage has the tendency of reducing the rate of hypotension, but apparently it does not reduce the incidence of bradycardia.