Effects of Leucosporidium-derived ice-binding protein (LeIBP) on bull semen cryopreservation.

We examined the effect of ice-binding protein derived from Leucosporidium (LeIBP) on the cryopreservation of bull semen and compared it with that derived from previously reported Antifreeze Protein III (AFPIII). Six concentrations of LeIBP (10-1  ~ 104  µg/ml) and AFPIII (10-1  ~ 104  µg/ml) were added to the bull semen extender, respectively. Sperm kinematic parameters were measured to examine sperm toxicity and cryopreserved sperm quality. Measures of antioxidant activity such as superoxide dismutase (SOD), reduced glutathione/oxidative glutathione (GSH/GSSG), and total antioxidant capacity (TAC) were analysed to identify the effect of LeIBP on sperm quality. In addition, sperm viability was analysed using a flow cytometer and fluorescence microscope by SYBR14/PI staining. The results showed that the LeIBP groups (0.1, 1 and 10 µg/ml) were less toxic, and the quality of the sperm were dramatically improved in the extenders containing 0.1 µg/ml LeIBP among concentrations of LeIBP and AFPIII. The SOD activity of LeIBP was greater than that of AFPIII and control. In addition, sperm viability was enhanced in the LeIBP-treated group. In summary, LeIBP is a useful cryoprotective adjuvant for bull sperm cryopreservation, and the most efficient concentration of LeIBP is 0.1 µg/ml.

Influence of nutritional deficiency on prognosis of renal cell carcinoma (RCC).

OBJECTIVE: To evaluate the prognosis of patients with renal cell carcinoma (RCC) by nutritional status defined by body mass index (BMI), serum albumin and cholesterol. PATIENTS AND METHODS: This study retrospectively enrolled 1437 patients who underwent radical nephrectomy (932) or partial nephrectomy (505) for RCC. We assigned nutritional status according to the presence of none or one nutritional risk factor (control group) and two or all three of the following nutritional risk factors (nutritional deficiency group). The nutritional factors and thresholds were preoperative albumin level (<3.5 g/dL), preoperative cholesterol level (<220 mg/dL), and preoperative BMI (<23 kg/m(2) ) RESULTS: The patients' mean (sd) age was 55.23 (12.41) years and BMI was 24.36 (3.17) kg/m(2) . The mean (sd) serum cholesterol level was 180.07 (38.24) mg/dL, and the albumin level was 4.2 (0.45) g/dL. In all, 141 (9.8%) patients had none of the nutritional deficiency criteria, 802 (55.8%) had one, 429 (29.9%) had two, and 65 (4.5%) had all three. Clinicopathological variables, i.e. female gender, high tumour stage, positive lymph node metastasis, positive distant metastasis, high nuclear grade and non-clear cell type histopathology were associated with the nutritional deficiency group. In multivariate Cox analysis, nutritional deficiency was an independent predictor for RCC recurrence (hazard ratio [HR] 1.39, 95% confidence interval [CI] 1.05-1.83, P = 0.020) and RCC-related mortality (HR 2.06, 95% CI 1.39-3.03, P < 0.001). CONCLUSION: Nutritional deficiency defined by BMI, serum albumin and cholesterol is an important factor that predicts postoperative prognosis of patients with RCC who have undergone radical or partial nephrectomy.

Dissolving microneedles for transdermal drug delivery.

Microfabrication technology has been adapted to produce micron-scale needles as a safer and painless alternative to hypodermic needle injection, especially for protein biotherapeutics and vaccines. This study presents a design that encapsulates molecules within microneedles that dissolve within the skin for bolus or sustained delivery and leave behind no biohazardous sharp medical waste. A fabrication process was developed based on casting a viscous aqueous solution during centrifugation to fill a micro-fabricated mold with biocompatible carboxymethylcellulose or amylopectin formulations. This process encapsulated sulforhodamine B, bovine serum albumin, and lysozyme; lysozyme was shown to retain full enzymatic activity after encapsulation and to remain 96% active after storage for 2 months at room temperature. Microneedles were also shown to be strong enough to insert into cadaver skin and then to dissolve within minutes. Bolus delivery was achieved by encapsulating molecules just within microneedle shafts. For the first time, sustained delivery over hours to days was achieved by encapsulating molecules within the microneedle backing, which served as a controlled release reservoir that delivered molecules by a combination of swelling the backing with interstitial fluid drawn out of the skin and molecule diffusion into the skin via channels formed by dissolved microneedles. We conclude that dissolving microneedles can be designed to gently encapsulate molecules, insert into skin, and enable bolus or sustained release delivery.

Role of metabolism by flavin-containing monooxygenase in thioacetamide-induced immunosuppression.

Thioacetamide has been known to cause immune suppression. The object of the present study is to investigate the role of metabolic activation by flavin-containing monooxygenases (FMO) in thioacetamide-induced immune response. To determine whether the metabolites of thioacetamide produced by FMO causes the immunosuppression, methimazole, an FMO inhibitor, was used to block the FMO pathway. Antibody-forming cell (AFC) response measured in BALB/c mice sensitized with sheep red blood cells (SRBCs) was compared between the groups treated with thioacetamide in the presence or absence of methimazole pretreatment. The pretreatment abolished the decrease in AFC number observed in the mice treated with thioacetamide alone. In addition, when spleen cells isolated from untreated mice were exposed to thioacetamide with a drug-metabolizing system, liver microsome and NADPH, for 4 h in vitro prior to the stimulation with mitogens, such as lipopolysaccharide (LPS) or concanavalin A (Con A), spleen cell proliferation was also decreased. The inhibitory effect of thioacetamide on cell growth was not detectable without the liver microsome. Moreover, the thioacetamide-suppressed proliferation of spleen cells in the presence of the metabolic activation system was prevented when coincubated with either SKF-525A, a cytochrome P450 (P450) inhibitor, or methimazole. We also found that the level of interleukin-2 (IL-2) in the culture supernatant was decreased by thioacetamide treatment and that the decrease of IL-2 level can be prevented by either SKF-525A or methimazole coincubation. Since IL-2 is one of the responsible factors that determine the proliferation level of lymphocytes, the change of IL-2 production was consistent with that of lymphoproliferation. In conclusion, thioacetamide-induced immunosuppression was, at least in part, due to the metabolites produced by FMO as well as by P450.