RESUMEN

This document represents the first collaboration between two organizations, American Society of Parenteral and Enteral Nutrition and the Society of Critical Care Medicine, to describe best practices in nutrition therapy in critically ill children. The target of these guidelines is intended to be the pediatric (> 1 mo and < 18 yr) critically ill patient expected to require a length of stay greater than 2 or 3 days in a PICU admitting medical, surgical, and cardiac patients. In total, 2,032 citations were scanned for relevance. The PubMed/Medline search resulted in 960 citations for clinical trials and 925 citations for cohort studies. The EMBASE search for clinical trials culled 1,661 citations. In total, the search for clinical trials yielded 1,107 citations, whereas the cohort search yielded 925. After careful review, 16 randomized controlled trials and 37 cohort studies appeared to answer one of the eight preidentified question groups for this guideline. We used the Grading of Recommendations, Assessment, Development and Evaluation criteria to adjust the evidence grade based on assessment of the quality of study design and execution. These guidelines are not intended for neonates or adult patients. The guidelines reiterate the importance of nutritional assessment, particularly the detection of malnourished patients who are most vulnerable and therefore potentially may benefit from timely intervention. There is a need for renewed focus on accurate estimation of energy needs and attention to optimizing protein intake. Indirect calorimetry, where feasible, and cautious use of estimating equations and increased surveillance for unintended caloric underfeeding and overfeeding are recommended. Optimal protein intake and its correlation with clinical outcomes are areas of great interest. The optimal route and timing of nutrient delivery is an area of intense debate and investigations. Enteral nutrition remains the preferred route for nutrient delivery. Several strategies to optimize enteral nutrition during critical illness have emerged. The role of supplemental parenteral nutrition has been highlighted, and a delayed approach appears to be beneficial. Immunonutrition cannot be currently recommended. Overall, the pediatric critical care population is heterogeneous, and a nuanced approach to individualizing nutrition support with the aim of improving clinical outcomes is necessary.

Asunto(s)

RESUMEN

Nutrition has a coadjuvant role in the management of children with acute diseases. We aimed to examine nutritional status, macronutrient requirements and actual macronutrient delivery in bronchiolitis. The nutritional status was classified according to WHO criteria and resting energy expenditure (MREE) was measured using an indirect calorimeter. Bland-Altman analysis was used to examine the agreement between MREE and estimated energy expenditure (EEE) with standard equations. Based on the ratio MREE/EEE in relation to Schofield equation on admission, we defined the subjects' metabolic status. A total of 35 patients were enrolled and 46% were malnourished on admission, and 25.8% were hypermetabolic, 37.1% hypometabolic and 37.1% normometabolic. We performed a 24-h recall in 10 children and 80% were overfed (AEI: MREE >120%). Mean bias (limits of agreement) with MREE was 8.9 (-73.9 to 91.8%) for Schofield; 61.0 (-41 to 163%) for Harris-Benedict; and 9.9 (-74.4 to 94.2%) for FAO-WHO equation. Metabolism of infants with bronchiolitis is not accurately estimated by equations.

Asunto(s)

RESUMEN

Calcitonin has been approved for the treatment of osteoporosis and other diseases involving accelerated bone turnover for approximately 25 years. The most commonly studied and prescribed form is salmon calcitonin, which has a greater efficacy in clinical use. A wealth of well-controlled clinical studies have demonstrated that calcitonin preserves or increases bone mineral density (BMD) and reduces the risk of vertebral fractures in osteoporosis. Recent studies have indicated that while a low BMD is correlated with an increase in fracture risk, increases in BMD alone do not explain the antifracture efficacy of antiresorptive therapies such as calcitonin. Therapies that moderately increase BMD may reduce fracture risk by reducing the rate of bone turnover and maintaining the integrity of the trabecular architecture, resulting in the preservation of bone strength and quality in osteoporotic patients. An advantage of calcitonin that is not shared by other antiresorptive therapies is its direct analgesic effect on bone pain. Calcitonin has been demonstrated to be clinically useful in improving pain, not only from the acute vertebral fractures of osteoporosis, but also in Paget's disease, bone malignancies, and other sources of musculoskeletal pain. Drugs containing calcitonin may be approved for additional indications in the near future, and as more convenient routes of administration such as the oral route become available, the demand for the calcitonin peptide is expected to increase.

Asunto(s)

Asunto(s)

RESUMEN

An infant with symptomatic congenital Cytomegalovirus infection is reported. After the detection of abnormalities on cranial ultrasound scanning, magnetic resonance imaging of the brain revealed a complete absence of corpus callosum with a midline anterior tubulonodular lipoma. A proposed causative link between early in utero Cytomegalovirus infection and lipoma with agenesis of corpus callosum is discussed.

Asunto(s)

Asunto(s)

RESUMEN

A rat alpha-amidating enzyme (alpha-AE) cDNA has been expressed in mouse C127 cells using a bovine papilloma virus vector in which transcription was regulated by the mouse metallothionein 1 promoter. The cDNA encoding the full length alpha-AE protein was modified to terminate translation at a site preceding the transmembrane and cytoplasmic domains, thereby enabling functional enzyme to be secreted into the medium. Purification of recombinant alpha-AE to homogeneity indicated that the enzyme was synthesized and secreted as two proteins of 75-77 kDA. The observed heterogeneity was due to inefficient glycosylation at Asn660, as demonstrated by glycopeptidase F digestion. Using the synthetic peptide, dansyl-Tyr-Val-Gly, the specific activity of the recombinant enzyme at pH 7.0 was found to be 1.4 mumol/min/mg and the Km of the enzyme was determined to be 3 microM. The purified recombinant enzyme has maximal activity at pH 4.5-5.5; however, a rapid inactivation of the enzyme occurs in acidic solutions in vitro. This inactivation is diminished when activity is measured at pH 7.0-10.0. The availability of large amounts of readily purified, active recombinant alpha-AE should allow detailed probing of reaction mechanism, copper coordination chemistry, and turn-over-based inactivation events.

Asunto(s)

Asunto(s)

RESUMEN

The alpha-amidating enzyme activity in rat medullary thyroid carcinoma (MTC) consists of multiple, active enzymes that can be resolved by ion-exchange chromatography. Amino acid sequences from one form of purified rat MTC alpha-amidating enzyme have been utilized to design oligonucleotide probes for isolating cDNAs encoding this protein. Sequence analysis of multiple cDNA clones indicates that there are at least two types of cDNA in rat tissues. These cDNAs differ primarily by the absence (type A) or the presence (type B) of a 315-base internal sequence. Additional heterogeneity in the 3' coding regions of the different mRNAs has also been found. Both types of cDNA predict primary translation products that are preproenzymes which must be post-translationally processed at both their amino and carboxyl termini. Sequence analysis of the purified peak III protein from rat MTC demonstrates that the type A mRNA encodes this 75-kDa protein. This analysis also provides support for the assignment of the C-terminal processing site. In addition, data are presented which demonstrate that type B mRNA is also functional. The implications of the internal and carboxyl-end heterogeneity are discussed.

Asunto(s)

RESUMEN

Conditioned medium from cultures of rat medullary thyroid carcinoma CA-77 cells was used as a source for purification of the secreted forms of peptidyl alpha-amidating enzyme. The alpha-amidating enzyme activity was partially purified using a combination of weak anion exchange and gel filtration chromatography. Subsequent strong anion exchange chromatography at pH 6.0 resolved this partially pure enzyme into four distinct peaks of activity, termed Ia, Ib, II, and III. Peaks Ia and Ib exhibited broad pH optima between pH 6.0-8.5, whereas peaks II and III both exhibited pH optima at approximately pH 5.0. The peak III activity was further purified to electrophoretic homogeneity using hydrophobic interactive chromatography followed by strong anion exchange chromatography at pH 8.0. The enzyme exhibited an apparent molecular mass of 75K, a pH optimum of approximately pH 5.0, and a maximal turnover number of 580 min-1 in the presence of L-ascorbate. Kinetic studies demonstrated that the enzyme probably functions through a ping-pong mechanism with respect to the binding of the glycine-extended peptide substrate and the L-ascorbate cofactor. The peak III enzyme exhibits several distinctive characteristics compared to amidating enzymes isolated and characterized by other laboratories.

Asunto(s)

Asunto(s)

RESUMEN

A peptidyl glycine alpha-amidating activity has been isolated from total tissue extracts of rat medullary thyroid carcinoma (MTC). Purification of the activity by ammonium sulfate fractionation, Sephacryl S-300 chromatography, and strong anion-exchange chromatography at pH 6.0 has resolved at least four peaks of activity. The activity associated with peak III has been further purified to apparent homogeneity by strong anion-exchange chromatography at pH 8.0. The purified peak III enzyme has an apparent molecular mass of 75,000 Da as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The identity of the 75,000-Da band as the alpha-amidating enzyme has been confirmed by recovery of activity from a nondenaturing polyacrylamide gel. The enzyme is catalytically active as a monomer, exhibits a pH optimum between 5.0 and 5.5, and has a turnover number of 300 min-1 for N-dansyl-Tyr-Val-Gly amidation at pH 5.5. The larger size, more acidic pH optimum, and higher specific activity distinguish the purified peak III rat MTC enzyme from the enzymes isolated from bovine and porcine pituitary or from frog skin.

Asunto(s)

RESUMEN

Potentially neoplastic outgrowths of BALB/c mouse mammary hyperplasia(s) (MH) derived from 7,12-dimethylbenz[a]anthracene-transformed epithelial cells in organ culture were characterized for responses to the hormones normally required for morphogenesis and functional differentiation of the mammary tissue. In a new culture model of the gland-free mammary fat pad, DNA synthesis in the MH outgrowths increased to a single peak during 6 days of incubation in a serum-free medium containing insulin, prolactin, aldosterone, and cortisol. The rise in DNA synthesis was accompanied by increased cell number and lobuloalveolar morphogenesis filling 50-70% of the fat pad; the rate of growth was variable among the MH outgrowth lines. Certain MH tissue showed some growth response in medium with aldosterone, cortisol, or estrogen, and progesterone in the presence of insulin. The hormone mixture insulin-prolactin was conducive to maximal growth, which suggested an altered sensitivity of the MH outgrowths to the mammogenic steroid hormones. Functional differentiation in the MH outgrowths was assessed in vivo by determination of the casein messenger RNA (mRNAcsn) levels measured by a specific complementary DNA probe. In MH-1 and MH-5 outgrowth lines, mRNAcsn was measurable in lactating hosts. In MH-9 outgrowth essentially the same concentration of mRNAcsn was present both in virgin and lactating hosts. Although mRNAcsn was present in MH outgrowths in lactating hosts, the concentration of the mRNA was only 0.056-0.88% of that present in the lactating host's own mammary gland. Virtually no mRNAcsn was measurable in the mammary tumors, regardless of the endocrine environment of the host animal. The results indicate a transformation-associated altered pattern of mammary cell-specific gene expression.

Asunto(s)

RESUMEN

Casein messenger RNAs (mRNAcsn) were purified from lactating mammary glands of BALB/c mice and used as a starting material for cloning of casein gene sequences. Double-stranded casein cDNA (ds-cDNAcsn) was prepared and blunt-end ligated to HindIII-specific DNA linker molecules. After digestion with HindIII, the dsDNAcsn was inserted into the HindIII site of plasmid pBR322, using T4 DNA ligase. Escherichia coli strain RH202 was transformed with the hybrid plasmids, and transformants were selected for resistance to ampicillin. Electrophoresis of HindIII-digested hybrid plasmid DNAs, followed by Southern transfer and hybridization to [32P]cDNAcsn, revealed that one of the hybrid-plasmid-containing colonies, designated pCas51, contained a 400-bp insert which hybridized to the [32P]cDNAcsn. Purification of the individual casein mRNAs (mRNAcsn alpha, beta, and gamma) and solution hybridization of nick-translated insert DNA to each of these revealed that pCas51 contained sequences complementary primarily to mRNAcsn beta.

Asunto(s)

RESUMEN

Entire second thoracic mammary glands of estrogen- and progesterone-treated immature virgin BALB/c mice were stimulated to pregnancylike lobuloalveolar morphogenesis after 6 days of incubation with insulin (5 micrograms/ml), aldosterone (1 micrograms/ml), growth hormone (5 micrograms/ml), cortisol (5 micrograms/ml), and prolactin (80 ng/ml, present as a contaminant in 5 micrograms/ml growth hormone). The alveolar growth in the glands, as judged by morphological studies, was accompanied by an increase in cell number as a function of incubation time in the hormonal medium. Hybridization of the total RNA from these glands to the casein mRNA specific complementary DNA probe (cDNAcsn) revealed that the level of casein mRNA rises from 0.00012 to 0.005% between 1 and 6 days of incubation. Estimates showed that the concentration of casein mRNA per cell rises 17-fold from 70 molecules on Day 1 to 1200 molecules on Day 6, whereas the number of epithelial cells increases only twofold during the same incubation time. When the growth hormone preparation was totally replaced by 80 ng of prolactin during the 6-day incubation, casein-mRNA levels were found to be 0.0083%. These results demonstrate that a pregnancy-like morphogenesis and concurrent expression of the casein gene in vitro can be achieved in a controlled hormone environment containing high cortisol and low prolactin concentrations. This one-step mammogenesis-lactogenesis culture model should be useful for studying the mechanisms of hormonal regulation of casein-gene expression observed in prepartum mammary gland in vivo.

Asunto(s)

RESUMEN

Second thoracic mammary glands of immature BALB/c female mice were stimulated to pregnancy-like lobuloalveolar (LA) development aftr 6 days of incubation in a corticosteroid-free step I culture medium containing insulin, prolactin, estradiol, progesterone, and growth hormone. A low basal level (0.0009%) of casein mRNA (mRNAcsn) sequences was detectable in the LA glands by a specific cDNA probe. Subsequent incubation of the LA glands for 3 days in medium containing insulin and prolactin or insulin and cortisol failed to elicit mRNAcsn above the basal level, indicating that neither prolactin nor cortisol alone can support casein gene expression. However, an increase in mRNAcsn levels was observed when the 3-day incubation with insulin and cortisol or insulin and prolactin was followed by 3 days of culture in presence of insulin, prolactin, and cortisol. When a 3-day incubation with insulin and prolactin was followed by 3 days in insulin and cortisol medium, mRNAcsn levels in the gland remained similar to the basal level. However, a 20-fold increase in the mRNAcsn levels ensued when the LA glands were sequentially incubated for 3 days in insulin and cortisol and then for another 3 days in insulin and prolactin medium. After a preincubation in insulin and cortisol medium, the LA glands retained residual cortisol during subsequent incubation in insulin and prolactin medium, and the mRNAcsn levels in these glands were related to the level of residual cortisol present. When mRNAcsn and the residual cortisol level reached a minimum, addition of fresh cortisol to the medium caused a 20-fold increase in the mRNAcsn levels. This indicates that cortisol is a limiting factor in insulin and prolactin medium and its presence is absolutely required for casein gene expression.

Asunto(s)

Asunto(s)

RESUMEN

The influence of cortisol and prolactin on casein gene expression in the mammary gland of lactating BALB/c mice was measured by using a specific cDNA probe to 15S casein mRNA (cDNAcsn). Casein mRNA (mRNAcsn) level in the mammary gland was decreased by 85% 5 days after adrenal ablation, but then was increased 4.4-fold 12 hr after a single injection of hydrocortisone-21-acetate. An 80% decrease in serum prolactin level, induced by the prolactin inhibitor 2-bromo-alpha-ergocryptin (CB-154), did not alter the level of mRNAcsn in the gland. Specific transcription of the casein gene in nuclei isolated from lactating mammary glands was measured by cDNAcsn hybridization to the in vitro synthesized Hg-CTP-containing RNA (Hg-RNA), which was purified by SH-agarose chromatography. The level of the mRNAcsn in Hg-RNA synthesized in the isolated nuclei was 0.09% and this was decreased 85% by alpha-amanitin, indicating that the mRNAcsn sequences in the Hg-RNA were the products of RNA polymerase II-directed DNA-dependent RNA synthesis. Transcription of the mRNAcsn in isolated nuclei was decreased by 70% 5 days after adrenalectomy and a single injection of the glucocorticoid then increased the transcription level 2-fold at 6 hr. Essentially no alteration of the level of transcription was detectable in mammary nuclei isolated from lactating mice with 80% decreased serum prolactin level, induced by CB-154 treatment. The results thus demonstrate a glucocorticoid involvement on the modulation of casein gene expression at the transcriptional level of control.

Asunto(s)

Asunto(s)

Asunto(s)