[Effects of irisin on lipid in mice and glycolipid metabolism in obese mice induced by diet].

Objective: To explore the role of low-dose irisin in the browning of white adipose tissue (WAT) and activation of brown adipose tissue (BAT) in mice, and its effect on the metabolic function of diet induced obesity. Methods: A total of 22 C57/BLKS/J male mice fed with normal diet and 8 fed with high fat diet were separately divided into experimental and control group. The experimental group was given irisin (0.8 ng/g, 200 µl), while the control group was given the same volume (200 µl) of phosphate buffer saline every day for 14 consecutive days intraperitoneally. Food intake and body weight of mice were collected regularly every day. After intervention, the mice were killed and the changes of lipid content and activity in adipose tissue were detected by histopathology and immunohistochemistry. The effects of irisin at different concentrations (0, 20 and 40 nmol/L) on primary white adipocytes and brown adipocytes were evaluated by immunohistochemistry on uncoupling protein 1(UCP1). In order to further evaluate whether irisin has the function of improving metabolism, the changes of serum indexes and hepatic steatosis in mice fed with high-fat diet were monitored. Results: The primary white and brown adipocytes derived from mice were successfully cultured and identified in vitro. In NCD mice, the weight gain of mice with irisin was lower than that of control mice [(-0.78±0.98) vs (0.27±0.55) g]. Histopathology showed that the area of white adipocytes with irisin was smaller than controls [(14.78±8.44) vs (29.49±12.97) µm2] and the brown adipocytes were larger than controls [(0.92±0.35) vs (0.19±0.12) µm2] (both P<0.05), while the expression of UCP1 in both adipose tissues was significantly higher in irisin group. After irisin treatment, the levels of blood glucose [(7.18±0.41) vs (13.48±2.07) mmol/L, P<0.01]and cholesterol [(2.38±0.26) vs (3.89±0.93) mmol/L, P<0.05] were significantly lower than controls, and the content of lipid droplets in liver cells was less than controls [ (2.73±1.96)% vs (14.04±6.29) %, P<0.001]. Conclusions: Low dose irisin can promote the browning of WAT and activate BAT, reducing the body weight of mice by producing heat. Irisin can also effectively improve diet-induced obesity and related metabolic disorders in mice.

[The value of Clermont score in detection intestinal mucosal ulcer of Crohn's disease].

Objective: To explore the value of Clermont score in the detection of intestinal mucosal ulcer in Crohn's disease (CD). Methods: From May 2015 to August 2017, 45 patients (28 males and 17 females; median age was 25 years) were confirmed to have ileocolic CD by endoscopic and pathological examinations at Nanjing General Hospital. All patients underwent MRE and DWI examinations. Based on the appearance of intestinal mucosa endoscopically, intestine segments from 45 patients were divided into three groups, namely, no ulceration group (NU), superficial ulceration group (SU), and deep ulceration group (DU). Several factors contribute to Clermont score calculation. These included the measurement of intestinal wall thickness using MRE, evaluation of intestinal wall edema and ulceration by MRE, DWI performance and ADC value of each segment. One-way ANOVA was utilized to compare the Clermont scores and ADC values of the intestine segments among the three groups. The efficacy of the Clermont scores and ADC values in evaluating intestinal mucosal ulcer in CD was verified using ROC analysis. Results: A total of 137 intestine segments were included in the study with 30 cases in NU, 37 cases in SU, and 70 cases in DU.DU had the highest Clermont score (22.5±4.5),following were SU(15.8±3.5) and NU(10.2±1.3)(F=179.935,P<0.01).The ADC values of DU ((1.34±0.17)×10(-3)mm(2)/s) was lower than NU ((2.07±0.52)×10(-3)mm(2)/s) and SU ((1.52±0.23)×10(-3) mm(2)/s) (F=83.822,P<0.01).The AUCs of using Clermont score and ADC value in differentiating deep ulcerations were 0.887 and 0.733, respectively. Conclusions: Either Clermont score or ADC value can be used to evaluate mucosal ulcer in CD. Clermont score demonstrates a better efficacy than ADC value in detecting deep ulcerations.

[Assessment of renal function with intravoxel incoherent motion and diffusion tensor imaging in type 2 diabetic patients].

Objective: To evaluate the application value of intravoxel incoherent motion (IVIM) and diffusion tensor imaging (DTI) in detecting early-stage diabetic nephropathy and to assess the damage of ralated renal function. Methods: A total of 52 patients with type 2 diabetes diagnosed in Zhongda Hospital were collected from April 2016 to May 2017 and were assigned to DM group (diabetes without nephropathy, n=32) and DN group (diabetes with nephropathy, n=20) according to detection of microalbuminuria, a cohort of healthy recipients were included as control group (n=27) in the meantime. All of the subjects underwent IVIM and DTI examination. The cortical and medullary parameters[IVIM: perfusion fraction f, tissue diffusivity D, pseudodiffuvisity D(*;) DTI: fractional anisotropy FA, apparent diffusion coefficient ADC, principal diffusivities (λ1, λ2, λ3)]were obtained respectively and were compared among groups. The relationship between MRI related parameters and estimated glomerular filtration rate (eGFR) were statistically investigated; and diagnostic performance of IVIM and DTI in discriminating DM and DN group was evaluated by receiver operating characteristic analysis. Results: The cortical and medullary f, D values in DN group were lower than those in DM group and control group (F=17.32, 15.69, 6.71, 10.94, all P<0.05). D values of all subjects showed positive correlations with eGFR (cortex r=0.518, medulla r=0.538, both P<0.05). The diagnostic efficiency of cortical f values to discriminate diabetes and diabetic nephropathy was 0.817, the cut-off value was 0.205. The medullary FA value in DM group was lower than that in control group ((0.371±0.051 vs 0.423±0.043, t=4.188, P<0.05); and the medullary FA value in DN group (0.315±0.062) was lower than that in control and DM group (F=25.08, P<0.05). The medullary λ3 values in DM group and DN group were all significantly higher than that in control group (F=7.86, P<0.05). The diagnostic efficiency of medullary FA values to discriminate diabetes and diabetic nephropathy was 0.763, the cut-off value was 0.344. Conclusion: IVIM and DTI can reflect the abnormal perfusion and diffusion during early-stage diabetic nephropathy and have the potential value to assess the damage of ralated renal function.

Cell therapy for salivary gland regeneration.

There are still no effective therapies for hyposalivation caused by irradiation. In our previous study, bone marrow stem cells can be transdifferentiated into acinar-like cells in vitro. Therefore, we hypothesized that transplantation with bone marrow stem cells or acinar-like cells may help functional regeneration of salivary glands. Bone marrow stem cells were labeled with nanoparticles and directly co-cultured with acinar cells to obtain labeled acinar-like cells. In total, 140 severely combined immune-deficiency mice were divided into 4 groups for cell therapy experiments: (1) normal mice, (2) mice receiving irradiation around their head-and-neck areas; (3) mice receiving irradiation and intra-gland transplantation with labeled stem cells; and (4) mice receiving irradiation and intra-gland transplantation with labeled acinar-like cells. Our results showed that salivary glands damaged due to irradiation can be rescued by cell therapy with either bone marrow stem cells or acinar-like cells for recovery of saliva production, body weight, and gland weight. Transdifferentiation of bone marrow stem cells into acinar-like cells in vivo was also noted. This study demonstrated that cell therapy with bone marrow stem cells or acinar-like cells can help functional regeneration of salivary glands, and that acinar-like cells showed better therapeutic potentials than those of bone marrow stem cells.

Normal pressure hydrocephalus after gamma knife radiosurgery for vestibular schwannoma.

Vestibular schwannomas are not uncommon, and gamma knife radiosurgery is one of the treatment options for symptomatic tumors. Hydrocephalus is a complication of gamma knife treatment of vestibular schwannoma, though the mechanism of the development of hydrocephalus remains controversial. We present an unusual case of normal pressure hydrocephalus (NPH) after gamma knife radiosurgery of a vestibular schwannoma in which the timeline of events strongly suggests that gamma knife played a contributory role in the development of the hydrocephalus. This is probably the first case of NPH post radiosurgery with normal cerebrospinal fluid protein. Communicating hydrocephalus should be treated with placement of shunt while non-communicating hydrocephalus can be treated with third ventriculostomy. Frequent monitoring and early intervention post radiosurgery is highly recommended to prevent irreversible cerebral damage.

Substitution of the critical methionine residues in trigonopsis variabilis D-amino acid oxidase with leucine enhances its resistance to hydrogen peroxide.

Each of the six oxidative-sensitive methionine residues in Trigonopsis variabilis D-amino acid oxidase (DAAO) was changed to leucine by site-directed mutagenesis. The wild-type and mutant enzymes with an apparent molecular mass of about 39.3 kDa were expressed in Escherichia coli. The specific activity of four mutant DAAOs (Met(104)Leu, Met(226)Leu, Met(245)Leu, and Met(339)Leu) was decreased by more than 96%, while Met(156)Leu and Met(209)Leu showed about 23% and 96% higher activity, respectively, than the wild-type enzyme. The kinetic parameters of the two more active enzymes were determined and a 2.2-fold increase in K(m) was observed for Met(209)Leu. Comparison of Met(156)Leu and wild-type DAAO revealed a 95% increase in k(cat)/K(m). Met(156)Leu, Met(209)Leu, and Met(226)Leu were resistant to inactivation by 50 mM H(2)O(2). The other three mutant DAAOs were also slightly more resistant than the wild-type enzyme to chemical oxidation. These observations indicate that the oxidative stability in T. variabilis DAAO can be improved by substitution of methionine residues with leucine.

The role of a conserved histidine residue, His324, in Trigonopsis variabilis D-amino acid oxidase.

To investigate the functional role of an invariant histidine residue in Trigonopsis variabilis D-amino acid oxidase (DAAO), a set of mutant enzymes with replacement of the histidine residue at position 324 was constructed and their enzymatic properties were examined. Wild-type and mutant enzymes have been purified to homogeneity using the His-bound column and the molecular masses were determined to be 39.2 kDa. Western blot analysis revealed that the in vivo synthesized mutant enzymes are immuno-identical with that of the wild-type DAAO. The His324Asn and His324Gln mutants displayed comparable enzymatic activity to that of the wild-type enzyme, while the other mutant DAAOs showed markedly decreased or no detectable activity. The mutants, His324/Asn/Gln/Ala/Tyr/Glu, exhibited 38-181% increase in Km and a 2-10-fold reduction in kcat/Km. Based on the crystal structure of a homologous protein, pig kidney DAAO, it is suggested that His324 might play a structural role for proper catalytic function of T. variabilis DAAO.

A conserved aspartate is essential for FAD binding and catalysis in the D-amino acid oxidase from Trigonopsis variabilis.

To evaluate the possible contribution of Asp206 of Trigonopsis variabilis D-amino acid oxidase (DAO) to its flavin adenine dinucleotide (FAD) binding and catalytic function, six mutant enzymes were constructed by site-directed mutagenesis. Western immunoblot analysis revealed that a protein with an apparent molecular mass of about 39.2 kDa was present in the cell-free extracts of wild-type and mutant strains. Replacement of Asp206 with Leu, Gly, and Asn resulted in the loss of DAO activity and characteristic absorption spectrum for flavoenzyme, while the other mutant DAOs, Asp206Glu, Asp206Ser, and Asp206Ala, exhibited a similar spectral profile to that of wild-type enzyme and retained about 6-90% of the enzyme activity. These results suggested that Asp206 of T. variahilis DAO might play an important role in the binding of FAD.

[Clinical assessment of desmopressin to reduce blood loss in patients after cardiopulmonary bypass].

Blood loss in patients on cardiopulmonary bypass is tremendously large after surgery due to platelet dysfunction. Desmopressin acetate (a synthetic analogue of vasopressin) has been shown to improve blood coagulation in a variety of platelet disorders especially in patients with hemophilia, von Willebrand's disease and uremia. Recent years, it has been used to treat patients with severe platelet dysfunction and profuse hemorrhage after cardiopulmonary bypass. We have investigated the effect of desmopressin acetate administration in randomized trials of 48 adult patients placed on cardiopulmonary bypass during cardiac surgery. Twenty four patients received intravenous infusion 0.3 microgram/kg desmopressin acetate one hour after cardiopulmonary bypass and other patients only received a placebo. Comparing with the control group, patients receiving desmopressin had shortened bleeding time (3.4 +/- 0.6 vs 5.1 +/- 1.6 mins, 8 hrs post bypass), lessened blood loss (482 +/- 258 ml vs 1430 +/- 733 ml, 24 hrs post bypass) and received fewer blood component therapy (pack RBC 2.7 +/- 2.2 vs 6.6 +/- 3.2 units, FFP 4.3 +/- 2.4 vs 11.7 +/- 5.7 units, platelet 3.8 +/- 5.0 vs 8.4 +/- 7.2 units). We conclude that desmopressin acetate can improve blood coagulation ability with safety and in reducing blood loss in patients after cardiopulmonary bypass.