Short communication: Recombinant mammary serum amyloid A3 as a potential strategy for preventing intramammary infections in dairy cows at dryoff.

Mammary serum amyloid A3 (M-SAA3) has shown potential in stimulating innate immunity during intramammary infections, at calving and at dryoff. In this study, we produced recombinant caprine M-SAA3 to test its ability to reduce intramammary infections with Staphylococcus aureus, Streptococcus uberis, Streptococcus dysgalactiae, and Escherichia coli, which are all common mastitis-producing pathogens. Recombinant production of M-SAA3 (followed by lipopolysaccharide removal to avoid lipopolysaccharide-nonspecific stimulation of the immune system) was successfully achieved. Mammary serum amyloid A3 stimulated the expression of IL-8 in a dose-dependent manner in primary mammary cultures. Although a direct killing effect on Staph. aureus by M-SAA3 was not detected, this acute phase protein was able to reduce Staph. aureus, Strep. uberis, and Strep. dysgalactiae infections by up to 50% and induced a reduction in E. coli counts of 67%. In general, the best concentration of caprine M-SAA3 for inhibiting infections was the lowest concentration tested (10 µg/mL), although higher concentrations (up to 160 µg/mL) increased its antimicrobial potential against some pathogens.

Cyclopropanation of enantiopure metal alkenyl carbenes with 2-methoxyfuran: a practical route to carboxycyclopropylglycine precursors.

We have examined the reactivity of enantiopure alkenyl Fischer carbene complexes 1, readily available from the chiral pool, with 2-methoxyfuran 4. In this reaction, polyfunctionalised cyclopropylcarbenes 5 are obtained under very mild conditions and with high selectivity, the major stereoisomer being isolated in an enantiopure form. The reaction involves the conjugate nucleophilic addition of 2-methoxyfuran 4 to the carbene complexes 1 followed by ring closure of the resulting zwitterionic intermediate species. The oxidation of the carbene 5 a results in the formation of the enantiopure cyclopropane diester 6. Further elaboration of the cyclopropane 6 allows for an efficient enantioselective access to alcohols or diols 7-9 as well as to cyclopropanecarbaldehydes 10-12. The protocol described herein provides a very simple entry to interesting enantiopure precursors of carboxycyclopropylglycine derivatives from readily available starting materials. In order to test this potential as carboxycyclopropylglycine precursors, the aminocyanation of the cyclopropanecarbaldehyde 10 was undertaken and the alpha-aminocyano derivative 13 was isolated as a single diastereosiomer.

Endothelial nitric oxide synthase gene polymorphism in dialysis patients.

Nitric oxide is an important factor in the regulation of vasodilator tone. In vascular cells, NO is synthesized by endothelial nitric oxide synthase, a key enzyme of the endogenous vasodilator system. Some studies have described the interaction between NO and the other factors that promote vasodilatation in vascular smooth muscular cells. Some of those factors are angiotensin-converting enzyme (ACE), transforming growth factor beta (TGF beta), and endothelial oxide nitric synthase (eNOS). Polymorphism that can alter the expression or the function of the eNOS protein has been identified in the eNOS gene in the promoter and codification zones. We studied the Glu298Asp variant of the eNOS gene in 52 hemodialysis (HD) patients, 22 peritoneal dialysis (PD) patients, and 93 healthy controls. Identification of the Glu298Asp variant in exon 7 was performed by enzymatic amplification and restriction fragment length polymorphism (RFLP) analysis. The frequencies of eNOS genotypes in the control group were GG, 39.8%; GT, 43%; and TT, 17.2%. In HD patients, the frequencies were GG, 40.3%; GT, 38.7%; and TT, 21.7%. In PD patients, they were GG, 41.6%; GT, 50%; and TT, 8.6%. No significant differences were seen between the control group and the dialysis patients, or between the HD and the PD patients.

Protein oxidative stress in dialysis patients.

Inflammatory status is observed in patients with chronic renal failure (CRF). The relationship between oxygen free radical production and dialysis could play an important role in protein oxidation. Carbonyl protein plasma level is an important tool in the study of protein stress, and it is related to the arterial intima thickness in the atherosclerosis process. We studied protein oxidative stress in 21 peritoneal dialysis (PD) patients and 42 hemodialysis (HD) patients as compared with 32 undialyzed patients with CRF. Carbonyl protein plasma levels were measured in nanomoles per milligram protein by the ELISA method (Winterbourn et al). Dialysis patients had a higher protein carbonyl content than did CRF patients (0.1265 +/- 0.04 nmol/mg vs. 0.1594 +/- 0.03 nmol/mg, p < 0.0002). Patients on PD had a lower level than patients on HD (0.1452 +/- 0.03 nmol/mg vs. 0.1665 +/- 0.04, p < 0.004). Glucose administration in PD is known to be able to increase glucose degradation products (GDPs) and advanced glycosylation end-products (AGEs) with high carboxylic and oxidative stress. In our study, the carbonyl protein level was higher in HD patients than in PD patients, perhaps because more protein oxidative stress is associated with hemodialysis technique or because the PD patients had greater residual renal function.