Leptin deficient ob/ob mice and diet-induced obese mice responded differently to Roux-en-Y bypass surgery.

OBJECTIVE: Weight regain contributes to the therapeutic failure in 15-20% of type 2 diabetic patients after Roux-en-Y gastric bypass surgery (RYGB), and the mechanism remains largely unknown. This study was conducted to explore the mechanism of weight regain. RESEARCH DESIGN: Wild-type (WT) diet-induced obese (DIO) mice were used to mimic human obesity, and ob/ob mice were used for leptin deficiency-induced obesity. Two groups of mice were compared in weight regain for 10 months after RYGB. Weight loss, food intake, fecal energy loss and energy expenditure were monitored in the study of weight regain. Fasting insulin, insulin tolerance and homeostatic model assessment-insulin resistance were tested for insulin sensitivity under the weight regain. Weight loss from RYGB and calorie restriction was compared for the impact in insulin sensitivity. RESULTS: In WT mice, RYGB induced a sustained weight loss and insulin sensitization over the sham operation in this 10-month study. However, RYGB failed to generate the same effects in leptin-deficient ob/ob mice, which suffered a weight regain over the pre-surgery level. In ob/ob mice, body weight was reduced by RYGB transiently in the first week, recovered in the second week and increased over the baseline thereafter. Weight loss was induced by RYGB relative to that of sham mice, but the loss was not sufficient to keep body weight below the pre-surgery levels. In addition, insulin sensitivity was not improved by the weight loss. The response to RYGB was improved in ob/ob mice by 2 weeks of leptin treatment. Weight loss from calorie restriction had an equivalent effect on insulin sensitization compared with that of RYGB. CONCLUSION: Those data demonstrate that ob/ob mice and DIO mice responded differently to RYGB surgery, suggesting that leptin may be one of the factors required for RYGB to prevent weight regain and diabetes recurrence.

Acute and long-term transcriptional responses in sulfur mustard-exposed SKH-1 hairless mouse skin.

Sulfur mustard (HD) ranks among the alkylating chemical warfare agents. Skin contact with HD produces an inflammatory response that evolves into separation at the epidermal-dermal junction conducting to blistering and epidermis necrosis. Up to now, current treatment strategies of HD burns have solely consisted in symptomatic management of skin damage. Therapeutic efficacy studies are still being conducted; classically using appropriate animal skin toxicity models. In order to substantiate the use of SKH-1 hairless mouse as an appropriate model for HD-induced skin lesions, we investigate the time-dependent quantitative gene expression of various selected transcripts associated to the dorsal skin exposure to HD saturated vapors. Using quantitative real time polymerase chain reaction (RT-qPCR), the expression of interleukins (IL-1ß and IL-6), tumor necrosis factor (TNF)-α, macrophage inflammatory proteins (MIP)-2α (also called Cxcl2) and MIP-1αR (also called Ccr1), matrix metalloproteases (MMP-9 and MMP-2), laminin γ2 monomer (Lamc2) and keratin (K)1 was determined up to 21 days after HD challenge in order to allow enough time for wound repair to begin. Specific transcript RT-qPCR analysis demonstrated that IL-6, IL-1ß, Ccr1, Cxcl2 mRNA levels increased as early as 6 h in HD-exposed skins and remained up-regulated over a 14-day period. Topical application of HD also significantly up-regulated MMP-9, TNF-α, and Lamc2 expression at specific time points. In contrast, MMP-2 mRNA levels remained unaffected by HD over the time-period considered, whereas that long-term study revealed that K1 mRNA level significantly increased only 21 days after HD challenge. Our study hereby provides first-hand evidence to substantiate a long period variation expression in the inflammatory cytokine, MMPs and structural components following cutaneous HD exposure in hairless mouse SKH-1. Our data credit the use of SKH-1 for investigating mechanisms of HD-induced skin toxicity and for the development of pharmacological countermeasures.

Penicillin acylase purification with the aid of hydrophobic charge induction chromatography.

The aim of this work was to test a chromatographic support, 4-mercaptoethyl pyridine (4-MEP) Hypercel, for penicillin acylase purification by using pure penicillin acylase and crude extract. Two equilibration buffers with various salt concentrations and different flow rates were tested. The relationships between electrostatic and hydrophobic interactions and proteins are demonstrated. (NH4)2SO4 proved preferable because no salting-in occurred, contrary to NaCl. The recovery and purification fold were similar to those obtained in pseudo-affinity chromatography with a three-fold reduction of the (NH4)2SO4 concentration.

Cloning, expression and two-step purification of recombinant His-tag enhanced green fluorescent protein over-expressed in Escherichia coli.

In this report, we describe a two-step chromatographic procedure for the purification of His-tag EGFP by immobilized metal affinity expanded bed adsorption (IMAEBA) as the capture step and size exclusion chromatography as the polishing step. The use of proteins including a histidine-tag facilitates their subsequent purification after expression in many microorganisms. This meets the needs of scientific researchers as well as industrialists in purifying recombinant proteins. The procedure described allowed the obtention of 230 mg pure EGFP from 1 l simple batch culture with a recovery of 90%.

Modification of flavonoid using lipase in non-conventional media: effect of the water content.

The enzymatic acylation of a flavonoid (naringin) was investigated in this work. This atypic substrate for a lipase was esterified very selectively by the immobilized Candida antarctica lipase: a single product was synthesized and was assumed to be the 6-O-palmitate naringin ester acylated on the glucose moiety. As lipase-catalyzed esterification reactions in organic media are greatly influenced by the water content, the effect of the initial hydration level of the reaction medium components was pointed out for naringin palmitate synthesis. 2-Methyl 2-butanol (solvent) and naringin (acyl acceptor) provided high amounts of water and when dried increased the conversion yield by 63% and the specific activity by 60%. On the contrary, the enzyme must not be dried because water is essential for the three-dimensional structure of the protein and, if absent, results in a 67% loss of activity. As water was produced in parallel to ester synthesis, the equilibrium of the reaction might be shifted by its removal. When the reaction was carried out with 100 g l(-1) molecular sieves 4A added after 24 h of reaction, a conversion yield of 43% was reached after 55 h reaction.

Enzymatic hydrolysis of (chloromethyldimethylsilyl)-2-propenyl acetate isomers: atypic specificity of Candida antarctica lipase.

Enzymatic hydrolysis of a mixture of (chloromethyldimethylsilyl)-2-propenyl acetate isomers was investigated by using immobilized Candida antarctica lipase as biocatalyst. TLC analysis and 1H NMR spectroscopy were used to monitor the extent of the reaction. At 60 degrees C, the enzyme exhibited a high selectivity towards 3-(chloromethyldimethylsilyl)-2-propenyl acetate which was almost quantitatively hydrolyzed, whereas, only 11% of 2-(chloromethyldimethylsilyl)-2-propenyl acetate reacted with the lipase. Consequently, the unreacted acetate was readily purified from the reaction medium by flash column chromatography and deacetoxylated in acidic methanol to give the corresponding hydroxy compound in a 71% global yield. On the other hand, without lipase, chemical treatment of the acetate mixture resulted in much lower yields in hydroxy compounds followed by a tedious purification process.

Construction of natural language sentence acceptors by a supervised-learning technique.

The problem discussed here is to build automatically an acceptor for natural language sentences, a sample of sentences being given. We solve it in the context of computer-assisted instruction and database interrogation. This paper is focused toward the definition of a learning criterion, i.e., a quality measure on the set of the acceptors which are compatible with the sample.