Evaluation of human milk fortification from the time of the first feeding: effects on infants of less than 31 weeks gestational age.

OBJECTIVE: To determine whether human milk fortification from the time of the first feeding significantly improves weight gain and bone mineral status in infants of <31 weeks estimated gestational age as compared with delayed or standard human milk fortification. STUDY DESIGN: This was a retrospective pre-post design. In all, 95 infants born at <31 weeks estimated gestational age were compared. There were 53 infants in the early fortification group (EFG) and 42 infants in the delayed fortification group (DFG). They were compared with regard to weight gain at 34 weeks postmenstrual age (PMA), and their serum levels of calcium, phosphorus and alkaline phosphatase levels were compared as an indicator of bone mineral status. The practice change of fortifying all human milk given to preterm infants at <34 weeks PMA commenced in June 2009. The usual practice of fortification took place once an infant had reached a feeding volume of 50 to 100 ml kg(-1) per day. The new practice fortified all human milk with a powdered human milk fortifier to 24 calories per ounce, starting with the first feeding, no matter how small the volume. RESULT: There were no differences in weight gain between the EFG and the DFG. The group that received fortification from the time of the first feeding were significantly less likely to have alkaline phosphatase levels >500 U l(-1) from 33 weeks PMA onward. There was no incidence of feeding intolerance with early fortification. CONCLUSION: Fortification of human milk from the time of the first feeding does not affect weight gain at 34 weeks PMA, but is related to a lower incidence of elevated alkaline phosphate levels and does not cause feeding intolerance.

Cleavage of double-stranded copy RNA by RNase 1 and RNase T1 provides a robust means to detect p53 gene mutations in clinical specimens.

Detecting somatic mutations in patient specimens is challenging because of the wide variation in quality and quantity of genomic DNA in clinically derived material. In cancer specimens, the challenge of detecting mutations is usually compounded by the presence of large numbers of nonmutated normal cells that dampen the relative signal that can be obtained from employing any mutation detection strategy. In the case of somatic mutations in the gene encoding the tumor suppressor, p53, a clinically useful mutation detection assay must be able to detect a wide variety of types of mutations scattered over five coding exons and their flanking intron sequences. This study examined the ability of a mutation detection strategy, termed NIRCA, to identify single-base mutations in the clinically relevant domain of the p53 gene. This strategy relies on RNase digestion-mediated cleavage of double-stranded copy RNA transcribed in vitro from polymerase chain reaction (PCR)-amplified genomic templates to detect mismatched base pairs resulting from hybridization of complimenting mutant and wild-type copy RNA strands. This assay system was found to robustly detect all twelve possible mismatches and the plus one and minus one frame shifts. Furthermore, the assay could detect mutations in clinical specimens when the mutant alleles composed as few as 4% of the total population of alleles isolated in bulk specimen genomic DNA. This mutation detection strategy worked efficiently in bladder, breast, colon and lung tumors as well as sediments from bladder cytology specimens.

Immunohistochemical localization of nitric oxide synthase in the autonomic innervation of the human penis.

An improved understanding of the physiology of penile erection has resulted from recent evidence that implicates nitric oxide as the principal mediator of erectile function. Previously, the neuroanatomy of erection in man was established with descriptions of the autonomic innervation of the pelvic organs and external genitalia. The basis upon which novel physiological concepts of erection relate to earlier neuroanatomical principles remains to be determined. In the present study these relationships were explored with nitric oxide synthase immunohistochemistry and reduced nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase histochemistry of select pelvic tissue specimens obtained from 4 men (3 at radical prostatectomy and 1 at autopsy). Nitric oxide synthase, the enzyme that catalyzes nitric oxide production, was identified in discrete neuronal locations, including the pelvic plexus, cavernous nerves and their terminal endings within the corporeal erectile tissue, branches of the dorsal penile nerves and nerve plexuses in the adventitia of the deep cavernous arteries. This distribution of nitric oxide synthase-containing nerves suggests that nitric oxide neuronally modulates local vascular smooth musculature of the penis. On this basis, nitric oxide is identified as a neuronal mediator of penile erection in man.

Reduction of epididymal sperm motility after ablation of the inferior mesenteric plexus in the rat.

Surgical ablation of the inferior mesenteric plexus in the rat results in an excessive accumulation of sperm within the cauda epididymidis. The present study examined the effect of ablation of the inferior mesenteric plexus on the motility of cauda epididymal spermatozoa. One to 8 weeks after ablation of the inferior mesenteric plexus, sperm curvilinear and straight line swimming velocities were reduced significantly compared with sperm curvilinear and straight line swimming velocities from sham-operated controls. Sperm swimming linearity also changed significantly versus controls after inferior mesenteric plexus ablation. Additional experiments using a vasectomy model suggested that the reductions in sperm curvilinear and straight swimming velocities after inferior mesenteric plexus ablation were not primarily the result of tubal obstruction. These data suggest that the sympathetic nervous system may influence epididymal sperm function.

Ablation of the inferior mesenteric plexus in the rat: alteration of sperm storage in the epididymis and vas deferens.

The involvement of the sympathetic nervous system in the transport and storage of spermatozoa in the male reproductive tract was examined by surgically ablating the inferior mesenteric plexus (IMP). One to eight weeks after ablation of the IMP, epididymal weight and the total number of spermatozoa present in the cauda epididymidis were significantly greater in IMP-ablated rats than in sham-operated rats. By contrast, the number of spermatozoa present in the initial segment of the vas deferens was significantly greater than in sham operated controls one week after IMP ablation but returned to control levels at two, four, six and eight weeks. Throughout the experiment, no differences were observed between IMP-ablated and control rats in the percentage of motile cauda epididymal spermatozoa, testicular weight, testicular sperm number or serum testosterone. These data demonstrate that the sympathetic nervous system differentially regulates sperm transport and storage in the male reproductive tract and suggest that the IMP may influence the epididymal maturation of spermatozoa.

The effect of sulfhydryl oxidation on the morphology of immature hamster epididymal spermatozoa induced to acquire motility in vitro.

Immotile spermatozoa from the caput epididymidis become progressively motile when incubated in medium containing theophylline, seminal plasma, and albumin. We previously reported that under these incubation conditions the spermatozoa induced to acquire motility exhibited a marked flagellar angularity, with the sperm head or midpiece bent 90-180 degrees towards the tail. In addition, we demonstrated that sperm flagellar bending did not occur when the sulfhydryl oxidant diamide was added to the motility induction medium. In the present study, we examined further the effect of sulfhydryl oxidation on the morphology and sulfhydryl content of immature caput spermatozoa induced to acquire motility in vitro. We found that flagellar bending was prevented and sperm flagellar straightness was maintained in a dose-dependent manner by diamide. Moreover, flow cytometric analysis of caput sperm sulfhydryls using the sulfhydryl reagent monobromobimane (mBBr) revealed that 1) diamide oxidizes caput sperm sulfhydryls, and 2) less than 15% of the total reactive sperm sulfhydryls were oxidized at diamide concentrations capable of preventing sperm angulation. Sodium tetrathionate (NaTT), another sulfhydryl oxidant, and hamster cauda epididymal fluid (CEF) containing sulfhydryl oxidase enzyme activity also maintained flagellar straightness in induced caput spermatozoa and oxidized sperm sulfhydryls. The flagellar straightness in caput spermatozoa treated with sulfhydryl oxidants, however, was temporary; with extended incubation, diamide- or CEF-treated spermatozoa exhibited flagellar bending. Additional studies showed that the flagellar straightness observed in sulfhydryl-oxidized spermatozoa was sustained when nitrofurantoin, an inhibitor of glutathione reductase, was included in the induction medium. Flow cytometric analysis of nitrofurantoin-treated spermatozoa showed that nitrofurantoin maintained the sperm disulfides formed by diamide and prevented the reduction of sperm disulfides back to sulfhydryls. Taken together, these studies demonstrate the significance of sulfhydryl oxidation in maintaining the morphology of immature caput epididymal spermatozoa induced to acquire motility in vitro and suggest that sulfhydryl oxidation may be important in the development of motility during sperm epididymal maturation in vivo.