[Investigation on the cold-chain temperature of vaccine in some areas of Zhejiang Province].

Objective: To investigation the situation of cold chain on vaccine in parts of Zhejiang Province and to provide recommendations for the management. Methods: From October to December, 2016, we each selected an immunization clinic in Cangnan County of Wenzhou, Yongkang City of Jinhua, Jianggan District of Hangzhou. Temperature recorder and vaccine viral monitor (VVM) labels were used to monitor the cold chain during all the storage and transportation process. In Jianggan District, we use optical density sensor to detected 20 VVM labels every time when the vaccine was stock in and out. Results: In total, 54 958 records were collected by temperature recording devices in all the three immunization clinic. 275 records exceeded the temperature limit required for store and transportation, of which 270 (98.2%) were above 8 ℃ and 5 (1.9%) were under 2 ℃. Excessive temperature exposure mainly occurred during the transportation (38.2%, n=105), followed by storage process in CDCs at different levels (26.2%, n=72), stock in and out (20.7%, n=50) and storage in the refrigerators in immunization clinics (14.9%, n=41). The average optical density difference between VVM labels and the reference circular decreased from 0.404 to 0.344 when the vaccines were delivered from the Zhejiang provincial CDC to immunization clinics. The color of VVMs did not significantly changed before use. Conclusions: The potential risk of vaccine cold chain in the monitoring sites is over-temperature. The weak links of cold chain management include the transportation, storage process, and stock in and out.

A thin-layer chromatography-bioautographic method for detecting dipeptidyl peptidase IV inhibitors in plants.

A thin-layer chromatography (TLC)-bioautographic method was developed with the aim to detect dipeptidyl peptidase IV (DPP IV) inhibitors from plant extracts. The basic principle of the method is that the enzyme (DPP IV) hydrolyzes substrate (Gly-Pro-p-nitroaniline) into p-nitroaniline (pNA), which diazotizes with sodium nitrite, and then reacts with N-(1-naphthyl) ethylenediamine dihydrochloride in turn to form a rose-red azo dye which provides a rose-red background on the TLC plates. The DPP IV inhibitors showed white spots on the background as they blocked enzymolysis of the substrate to produce pNA. The method was validated with respect to selectivity, sensitivity, linearity, precision, recovery, and stability after optimizing key parameters including plate type, time and temperature of incubation, concentration of substrate, enzyme and derivatization reagents, and absorption wavelength. The results showed good lineary within amounts over 0.01-0.1µg range for the positive control, diprotin A, with the coefficient of determination (r(2))=0.9668. The limits of detection (LOD) and quantification (LOQ) were 5 and 10ng, respectively. The recoveries ranged from 98.9% to 107.5%. The averages of the intra- and inter-plate reproducibility were in the range of 4.1-9.7% and 7.6-14.7%, respectively. Among the nine methanolic extracts of medicinal herbs screened for DPP IV inhibitors by the newly developed method, Peganum nigellastrum Bunge was found to have one white active spot, which was then isolated and identified as harmine. By spectrophotometric method, harmine hydrochloride was found to have DPP-IV inhibitory activity of 32.4% at 10mM comparing to that of 54.8% at 50µM for diprotin A.

Characterization of MUSTN1 gene and its relationship with skeletal muscle development at postnatal stages in Pekin ducks.

Musculoskeletal embryonic nuclear protein 1 (MUSTN1) gene is involved in myogenic fusion and differentiation in rats. We previously showed the differential expression of MUSTN1 in week (W) 2 and W6 breast muscles of Pekin ducks. In this study, we further investigated its molecular characteristics and expression profiles in different tissues at W7 and in breast and leg muscles at W1, W3, W5, W7, and W9. The relationship between muscle development and muscle fiber areas was also investigated. A 358-bp cDNA sequence was obtained. The coding sequence of duck MUSTN1 cDNA encoded a 78-amino acid sequence, which showed high similarity with those of other species (96% similarity with zebra finch and 94% with chicken). In addition, a 6435-bp genomic DNA sequence of MUSTN1 was obtained. In total, 231 transcription factor-binding sites were found in the promoter region, and many of these transcription factors were involved in the regulation of muscle development. MUSTN1 expression in breast muscle increased from W1 to W5 and then decreased at W9. In leg muscle, the expression increased from W1 to W3 and then decreased. The relative growth rates of breast and leg muscle fibers reached their peaks at W3-W5 and W1-W3, respectively. Since the greatest relative growth rates appeared at the highest expression levels of the MUSTN1 gene, it was thought to play roles in duck muscle development. Our findings would be helpful in understanding the molecular characteristics and functions of the MUSTN1 gene in breast muscle development of ducks.

Characterization of myostatin gene (MSTN) of Pekin duck and the association of its polymorphism with breast muscle traits.

Myostatin, encoded by the MSTN gene, is a negative regulator of muscle growth, and its expression level in muscle tissue is closely correlated with muscle growth and satellite cell proliferation. To identify the characteristics of the Pekin duck MSTN gene and the relationship between its polymorphism and breast muscle traits in Pekin duck, cDNA cloning and analysis and the expression pattern in breast muscle development and polymorphism were performed using molecular cloning, quantitative real-time reverse-transcription polymerase chain reaction, and molecular marker technology. The results showed that a 1320-bp sequence, including a 93-bp 5'-UTR, 1128-bp CDS, and 99- bp 3'-UTR, was obtained, and two alternative splicing isoforms were detected. The alternative splicing isoforms encoded 375- and 251-amino acid residues. The amino acid sequence of Pekin duck MSTN was similar to other vertebrates and exhibited the highest similarity to chicken. The expression pattern of MSTN in breast muscle tissue showed a tendency to increase, except for a slight decrease at 6 weeks. Three single nucleotide polymorphisms were found in the Pekin duck MSTN gene by cDNA sequencing from different individuals. The T129C had significant association with breast muscle thickness, and the T952C had significant association with the fossilia ossis mastodi length. This study reveals the molecular characteristics of the Pekin duck MSTN gene and the relationship of its polymorphism with breast muscle traits in Pekin duck. Therefore, it can provide some useful basic understanding of MSTN functions.

Developmental characteristics of pectoralis muscle in Pekin duck embryos.

To confirm the entire developmental process and transition point of embryonic Pekin duck pectoral muscle, and to investigate the association between pectoral muscle development and their regulating genes, anatomical and morphological analyses of embryonic Pekin duck skeletal muscles were performed, and the expression patterns of its regulating genes were investigated. The anatomical analysis revealed that body weight increased with age, while increases in pectoral muscle weight nearly ceased after the embryo was 20 days of hatching (E20). The developmental morphological characteristics of Pekin duck pectoral muscle at the embryonic stage showed that E20 was the transition point (from proliferation to fusion) of Pekin duck pectoral muscle. The expression patterns of MRF4, MyoG, and MSTN indicated that E19 or E20 was the fastest point of pectoral muscle development and the crucial transition for Pekin duck pectoral muscle development during the embryonic stage. Together, these findings imply that E20 is the crucial transition point (from proliferation to fusion) of Pekin duck pectoral muscle and that there is no muscle fiber hypertrophy after E20. Results of this study provide further understanding of the developmental process and transition point of Pekin duck pectoral muscle during the embryo stage.

Alanylglutamine dipeptide and growth hormone maintain PepT1-mediated transport in oxidatively stressed Caco-2 cells.

Reactive oxygen species (ROS) produced by gut mucosal cells during conditions such as inflammatory bowel disease (IBD) may impair mucosal repair and nutrient transport/absorptive function. Absorption of di- and tripeptides in the small intestine and colon is mediated by the H(+)-dependent transporter PepT1, but effects of oxidative stress on di- and tripeptide transport are unknown. We assessed whether exposure to hydrogen peroxide (H(2)O(2)) influences dipeptide transport in human colonic epithelial (Caco-2) cells. Uptake of [(14)C]glycylsarcosine (Gly-Sar) was used to evaluate PepT1-mediated dipeptide transport. Exposure to 1-5 mmol/L H(2)O(2) for 24 h caused a dose-dependent decrease in Gly-Sar transport, which was associated with decreased PepT1 transport velocity (V(max)). Treatment with alanylglutamine (Ala-Gln) or growth hormone (GH) did not alter Caco-2 Gly-Sar transport in the absence of H(2)O(2). However, both Ala-Gln and GH prevented the decrease in dipeptide transport observed with 1 mmol/L H(2)O(2) treatment. Ala-Gln, but not GH, maintained cellular glutathione and prevented the decrease in PepT1 protein expression. Thus, these agents should be further investigated as potential therapies to improve absorption of small peptides in disorders associated with oxidative injury to the gut mucosa.

Regulation of glutathione redox status in lung and liver by conditioning regimens and keratinocyte growth factor in murine allogeneic bone marrow transplantation.

BACKGROUND: Reactive oxygen species (ROS) and glutathione (GSH) depletion contribute to organ injury after bone marrow transplantation (BMT). Keratinocyte growth factor (KGF) ameliorates graft-versus-host disease (GVHD)-associated organ injury in murine BMT models. METHODS: B10.BR mice received total body irradiation (TBI; day -1) +/- cyclophosphamide (Cy; 120 mg/kg/day i.p., days -3 and -2), then were transplanted on day 0 with C57BL/6 bone marrow + spleen cells as a source of GVHD-causing T cells. KGF (5 mg/kg/day subcutaneously [s.c.]) or saline was given on days -6, -5, and -4. Lung and liver GSH and oxidized GSH disulfide (GSSG) levels were measured on days 0 and 5 and glutathione redox potential (Eh) calculated. Organ malondialdehyde (MDA) was determined on day 5 as an index of ROS-mediated lipid peroxidation. RESULTS: In lung, TBI+BMT oxidized GSH Eh and increased MDA. Cy further oxidized lung GSH Eh. In liver, neither BMT regimen altered GSH redox status or MDA. KGF prevented the decrease in lung GSH after TBI+Cy and decreased lung MDA after both TBI and TBI+Cy. KGF increased liver GSH levels and GSH Eh after TBI and GSH Eh after TBI+Cy. CONCLUSIONS: In murine allogeneic BMT, TBI oxidizes the lung GSH redox pool and Cy exacerbates this response by 5 days post-BMT. In contrast, liver GSH redox status is maintained under these experimental conditions. KGF treatment attenuates the Cy-induced decrease in lung GSH, decreases post-BMT lung lipid peroxidation, and improves liver GSH redox indices. KGF may have a therapeutic role to prevent or attenuate GSH depletion and ROS-mediated organ injury in BMT.

Growth factors regulation of rabbit sodium-dependent neutral amino acid transporter ATB0 and oligopeptide transporter 1 mRNAs expression after enteretomy.

BACKGROUND: Sucessful intestinal adaptation after massive enterectomy is dependent on increased efficiency of nutrient transport. However, midgut resection (MGR) in rabbits induces an initial decrease in sodium-dependent brush border neutral amino acid transport, whereas parenteral epidermal growth factor (EGF) and growth hormone (GH) reverse this downregulation. We investigated intestinal amino acid transporter B0 (ATB0) and oligopeptide transporter 1 (PEPT 1) mRNA expression after resection and in response to EGF and/or GH. METHODS: Rabbits underwent anesthesia alone (control) or proximal, midgut, and distal resections. Full-thickness intestine was harvested from all groups on postoperative day (POD) 7, and on POD 14 from control and MGR rabbits. A second group of MGR rabbits received EGF and/or GH for 7 days, beginning 7 days after resection. ATB0 and PEPT 1 mRNA levels were determined by Northern blot analysis. RESULTS: In control animals, ileal ATB0 mRNA abundance was three times higher than jejunal mRNA, whereas PEPT 1 mRNA expression was similar. By 7 and 14 days after MGR, jejunal ATB0 mRNA abundance was decreased by 50% vs control jejunum. A 50% decrease in jejunal PEPT 1 message was delayed until 14 days after MGR. Treatment with EGF plus GH did not alter ATB0 mRNA expression but doubled PEPT 1 mRNA in the jejunum. CONCLUSION: The site of resection, time postresection, and growth factors treatment differentially influence ATB0 and PEPT 1 mRNA expression. Enhanced sodium-dependent brush border neutral amino acid transport with GH plus EGF administration is independent of increased ATB0 mRNA expression in rabbit small intestine after enterectomy.

Enteral nutrition and keratinocyte growth factor regulate expression of glutathione-related enzyme messenger RNAs in rat intestine.

BACKGROUND: Malnutrition is associated with increased reactive oxygen species (ROS) formation and depletion of the critical antioxidant glutathione (GSH) in the intestine. The malnutrition-induced decrease in gut GSH levels is prevented by recombinant keratinocyte growth factor (KGF) administration. We investigated whether enzymes that are induced by oxidants and modulate tissue GSH supply are regulated by enteral nutrients or KGF at the messenger RNA (mRNA) level. METHODS: Adult rats were fasted for 3 days alone or fasted for 3 days then refed ad libitum. In a second model, rats were fasted for 3 days and then refed ad libitum or 25% of ad libitum intake with daily intraperitoneal saline or recombinant KGF (5 mg/kg/d) for 3 subsequent days. mRNA levels for gamma-glutamylcysteine synthetase (gamma-GCS), gamma-glutamyl transpeptidase (gamma-GT), glutathione-S-transferase Ya-subunit, gastrointestinal glutathione peroxidase (GI-GPx), and non-selenium-dependent glutathione peroxidase (ns-GPx) were determined in ileum and colon by ribonuclease protection assay. RESULTS: Fasting increased ileal gamma-GCS, ns-GPx, and glutathione-S-transferase mRNAs (by 36%, 165%, and 130% of controls) and decreased GI-GPx mRNA (to 55% of controls). In the colon, mRNAs for GSH-related enzymes were unchanged by fasting or refeeding. Prolonged enteral nutrient restriction (25% refeeding after a 3-day fast) increased gamma-GCS and glutathione-S-transferase mRNAs (by >270% of controls), decreased GI-GPx mRNA (to <50% of controls) in ileum and colon and increased ns-GPx mRNA (by 180%) in colon. KGF treatment increased ns-GPx mRNA in the ileum and colon and glutathione-S-transferase mRNA in the colon (by >200% of controls). CONCLUSIONS: Enteral nutrient intake regulates GSH-related enzyme mRNA levels in the intestine, which may contribute to the decrease in mucosal GSH during malnutrition. Increased ns-GPx and glutathione-S-transferase mRNA levels during malnutrition and with KGF administration may increase detoxifying functions in the gut under these conditions.

Heterogeneity of dermal deposition of eosinophil granule major basic protein in acute lesions of atopic dermatitis.

Deposition of eosinophil granule major basic protein (MBP) often occurs in acute and chronic lesions of atopic dermatitis, but it is not clear what the factors may be that are related to the MBP deposition in some skin lesions of the disease. The purpose of this study was to determine whether a personal or family history of respiratory atopy is related to the intensity of MBP deposition in acute lesions. We immunohistochemically stained biopsy specimens from acute, non-oozing indurated erythematous lesions of atopic dermatitis with BMK-13, a monoclonal antibody which recognizes MBP. The subjects were 40 adult patients with atopic dermatitis. Of the 40 patients, 22 had a personal history of respiratory atopy, 8 had a family history of respiratory atopy, and 10 had neither a personal nor a family history of respiratory atopy. Deposition of MBP was observed in the specimens from 24 (60%) of the 40 patients examined. Furthermore, there were great individual differences in the intensity of MBP deposition. A strong MBP deposition was often seen in specimens from patients with atopic dermatitis who had a personal or family history of respiratory atopy, but was absent in specimens from those patients with atopic dermatitis who had neither a personal nor a family history of respiratory atopy. We conclude that a strong MBP deposition seems to occur in acute lesions of those patients with atopic dermatitis who have a predisposition to respiratory atopy.

Regulation of keratinocyte growth factor (KGF) and KGF receptor mRNAs by nutrient intake and KGF administration in rat intestine.

The aim of this study was to investigate the regulation of keratinocyte growth factor (KGF) and KGF receptor mRNAs by diet and KGF treatment in rat intestine. Fasting for three days up-regulated KGF and KGF receptor mRNA levels in ileum and increased KGF receptor mRNA expression in colon. KGF and KGF receptor mRNA levels returned toward control values with ad libitum refeeding but remained elevated when refeeding was limited to 25% of ad libitum intake. KGF treatment during nutrient repletion did not alter intestinal KGF mRNA levels but increased KGF receptor mRNA abundance in ileum and colon. We conclude that the increase in KGF and KGF receptor mRNAs induced by malnutrition may be an adaptive response to attenuate gut mucosal atrophy in this setting. The gut-trophic effects of KGF treatment may be mediated, in part, by up-regulation of the KGF receptor mRNA in small bowel and colon.

Interactions between nutrients and peptide growth factors in intestinal growth, repair, and function.

Several lines of evidence demonstrate that general nutritional status, specific nutrients (eg, zinc, glutamine), and certain trophic growth factors (eg, growth hormone, insulin-like growth factor I, keratinocyte growth factor, and glucagon-like peptide-2) have important interactions relevant for intestinal growth and function. Adequate nutritional status is critical for endogenous growth factor synthesis in the gut and other tissues and is an important mediator of organ responsiveness to exogenous growth factor administration. Both endogenously synthesized and exogenously administered growth factors upregulate nutrient uptake and utilization by gut mucosa, skeletal muscle, and other organs. Emerging data from both animal and human studies indicate that combinations of selected growth factors and specific nutrients may improve the growth, adaptation, and repair of the intestinal mucosa. Additional studies to determine basic mechanisms of nutrient-growth factor interactions and the safety and efficacy of treatment with combinations of specific nutrients and recombinant growth factors are needed. Results of these investigations should define new methods for support of the intestinal tract during short bowel syndrome (SBS), catabolic illness, and malnutrition.

Keratinocyte growth factor enhances glutathione redox state in rat intestinal mucosa during nutritional repletion.

Malnutrition decreases tissue levels of glutathione (GSH), a major endogenous antioxidant that detoxifies reactive oxygen species and promotes cell growth. This study determined the effects of the gut trophic peptide keratinocyte growth factor (KGF) on intestinal mucosal GSH concentrations and redox state in malnourished rats. Adult rats were food-deprived for 3 d, then consumed food ad libitum or 25% of ad libitum intake for 3 d with daily intraperitoneal administration of saline or KGF (5 mg.kg-1.d-1). Mucosal GSH and glutathione disulfide (GSSG) concentrations, crypt depth and total mucosal height were measured in the jejunum, ileum and colon. In the 25% of ad libitum-refed, saline-treated group, mucosal GSH was lower in all gut tissues (42% in jejunum, 38% in ileum, and 57% in colon), and the GSH/GSSG ratio was lower in the jejunum and ileum compared to that in the ad libitum-refed controls. KGF treatment with ad libitum refeeding increased GSH/GSSG in the jejunum, ileum and colon. Furthermore, in 25% of ad libitum refeeding, KGF normalized jejunal, ileal and colonic mucosal GSH content and significantly increased the mucosal GSH/GSSG ratio relative to rats treated with saline. Increased crypt depth and total mucosal height induced by KGF and feeding could be explained in part by increased mucosal GSH content. KGF treatment improved gut mucosal glutathione redox state in malnourished, refed rats. These data provide evidence that gut trophic hormones and food intake may independently support gut mucosal glutathione antioxidant capacity during nutritional repletion.

Gut-trophic effects of keratinocyte growth factor in rat small intestine and colon during enteral refeeding.

BACKGROUND: Keratinocyte growth factor (KGF) induces proliferation of gut epithelium in rat models, but KGF-nutrient interactions have not been studied. An experimental model of fasting-induced gut atrophy followed by different levels of enteral refeeding was used to investigate the influence of nutrient availability on the gut-trophic effects of exogenous KGF. METHODS: After a 3-day fast, rats were enterally refed either ad libitum or at 25% of ad libitum intake for 3 subsequent days. Either intraperitoneal KGF (5 mg/kg/d) or saline was given in each dietary regimen. Wet weight, DNA, and protein content were measured as indices of full-thickness cellularity in duodenum, jejunum, ileum, and colon. Villus height in small bowel segments and crypt depth in all gut tissues were measured as specific indices of mucosal growth. RESULTS: Refeeding at 25% of ad libitum intake significantly decreased full-thickness cellularity and mucosal growth indices in duodenum, jejunum, and ileum. In the colon, only protein content fell significantly and crypt depth was maintained. KGF administration during 25% refeeding did not alter full-thickness indices in any small bowel segment or affect jejunal mucosal growth. In contrast, KGF normalized duodenal villus height (p < .01) and duodenal and ileal crypt depth (p < .05) only in the 25%-refed model. KGF significantly increased ileal villus height in both ad libitum and 25%-refed rats (by 43% and 48%, respectively, p < .05) and markedly increased colonic cellularity and mucosal crypt depth with both levels of refeeding (p < .01). CONCLUSIONS: Rat small bowel growth is more sensitive than colon to the level of enteral refeeding after a 3-day fast. KGF administration does not affect jejunal growth, but specifically prevents atrophy of duodenal and ileal mucosa during hypocaloric, hyponitrogenous refeeding. In ileum and colon, some KGF-mediated growth responses are independent of the level of enteral refeeding. Thus gut-trophic effects of KGF and KGF interactions with the level of nutrient intake are tissue-specific.

Comparison of the relative quantities of gamma-mRNAs and fetal hemoglobin in SS patients with different haplotypes.

We have studied the relative levels of gamma-mRNA [%gamma/(gamma + beta)], Ggamma- and Agamma-mRNAs [%Ggamma/(Ggamma + Agamma)], hemoglobin (Hb) F, and the Ggamma and Agamma chains in some 50 patients with sickle cell anemia (SS) and with different haplotypes. As expected, the Hb F levels varied greatly and were high in patients with the Saudi Arabian-Indian haplotype. Similarly, the Ggamma values varied greatly (from 19.5 to 76.5%) and depended on the haplotypes. A rare haplotype, named Mor, was found in 3 SS patients, 1 of whom was a homozygote Mor/Mor; this haplotype is associated with the lowest Ggamma value (19.5% in the homozygote) and with a C-->T mutation at position -202 of the Agamma promoter. The levels of gamma-mRNA roughly parallel those of Hb F, but older patients have increased levels of mRNA, which appears not to be efficiently translated into Hb F. Similar observation have been reported for other hemoglobinopathies such as deltabeta-thalassemia heterozygotes and Hb Lepore heterozygotes. The relative quantity of Ggamma-mRNA was closely related to that of the Ggamma chain in the 15 patients who were studied; the Ggamma- to Agamma-mRNA ratio did not change with age.

Epidermal growth factor and human growth hormone accelerate adaptation after massive enterectomy in an additive, nutrient-dependent, and site-specific fashion.

BACKGROUND: After massive enterectomy (ME), remnant intestine undergoes compensatory adaptation. Epidermal growth factor (EGF) and human growth hormone (hGH) have each been shown to enhance total length small intestine nutrient transport after ME. This study aims to determine the differential effects of EGF and hGH on proximal and distal small intestinal remnants after ME. METHODS: New Zealand white rabbits underwent 70% mid-jejunoileal resection. After 1 week, animals received hGH (0.2 mg/kg/day), EGF (1.5 micrograms/kg/hr), hGH + EGF, or vehicle (equal volume) for 7 days. Sodium-dependent uptake of glucose, glutamine, alanine, leucine, and arginine into brush border membrane vesicles was quantitated. Serum insulin-like growth factor-I concentrations as well as proximal and distal villus and microvillus heights were measured. IGF binding protein-3 and -4 mRNA expression was determined in full-thickness proximal and distal gut remnants. RESULTS: Concomitant hGH and EGF treatment up-regulates glucose (100%), glutamine (80%), and leucine (60%) transport in the proximal remnant; alanine (150%) and arginine (400%) transport in the distal remnant; and microvillus height (25% to 35%) both proximally and distally. Serum IGF-I levels and gross villus heights were not different among groups. CONCLUSIONS: Co-infusion of hGH and EGF accelerates intestinal adaptation after ME in an additive, nutrient-dependent, and site-specific fashion via enhanced nutrient transport as well as microvillus hypertrophy.

Hb Lepore-Baltimore (delta 68Leu-beta 84Thr) and Hb Lepore-Washington-Boston (delta 87Gln-beta IVS-II-8) in central Portugal and Spanish Alta Extremadura.

Hb Lepore is one of the most common abnormal haemoglobins in Caucasians in Central Portugal and in the Spanish Alta Extremadura (0.28% in a survey of school children). A group of 19 Portuguese and 14 Spanish Hb Lepore carriers (all unrelated) was characterised at the molecular level by the polymerase chain reaction, sequencing and restriction enzyme analysis. The Portuguese and one Spanish carrier were heterozygous for Hb Lepore-Baltimore, whereas all other Spanish subjects were Hb Lepore-Washington-Boston carriers. Sequencing of the Hb Lepore-Baltimore gene further established the crossover at delta 68-beta 84, a region two codons (CDs) shorter than that previously described and easily confirmed by digestion with MaeI and BanI. Data from haplotype analysis suggest that this crossover occurred as an independent event on the Iberian Peninsula. The haematological data were similar in both groups except for the levels of Hb F and the G gamma chain, which were significantly higher in the Hb Lepore-Baltimore heterozygotes. Quantification of the globin chains and the mRNA transcripts showed that the delta beta gene is transcribed at a higher level than the delta gene with levels of translation giving rise to 10%-15% of Hb Lepore. The different levels of Hb F observed in the two groups are the results of the higher transcription rate of the gamma genes in Hb Lepore-Baltimore heterozygotes and an apparently less efficient translation of G gamma genes in Hb Lepore-Washington-Boston heterozygotes.

Human growth hormone induces system B transport in short bowel syndrome.

BACKGROUND: After massive enterectomy, remnant intestine undergoes compensatory adaptation. A combination of human growth hormone (hGH) and a glutamine-enriched modified diet induces further adaptation in patients with short bowel syndrome (SBS) on long-term total parenteral nutrition. The specific actions of each component, however, are not well-defined. METHODS: New Zealand White rabbits were randomized to control, sham operation, or SBS (70% midjejunoileal resection) groups and treated with either hGH or saline. Sodium-dependent uptake of glucose, glutamine, alanine, leucine, and arginine into brush border membrane vesicles was quantitated. Serum insulin-like growth factor-I (IGF-I) levels were determined by immunoradiometric assay. Mucosal mRNA expression of IGF-I and IGF binding protein 4 (IGFBP-4) was evaluated by northern blot analysis using rat cDNA probes. RESULTS: Glutamine and leucine transports were 33 and 39% greater, respectively, in the hGH-treated versus saline-treated SBS group (P < 0.05), supporting induction of system B amino acid transport. This upregulation was due, in part, to an 88% increase in glutamine carrier capacity (Vmax) with no change in carrier affinity (Km). Both hGH treatment and SBS increased serum IGF-I levels without direct correlation with increased nutrient transport. IGFBP-4 mRNA expression in small bowel mucosa of saline-treated SBS animals was significantly greater than saline-treated unoperated control values. Mucosal IGFBP-4 mRNA was not significantly altered from control in the other study groups. IGF-I mRNA expression was not detected in mucosa, but weak hybridization was noted in rabbit liver. CONCLUSIONS: Human growth hormone accelerates early adaptation in SBS by upregulation of system B carrier capacity. Serum IGF-I levels and mucosal IGF-I and IGFBP-4 mRNA expression did not directly correlate with this enhanced nutrient transport, suggesting that hGH might exert its adaptive effects by mechanisms that are independent from the IGF system in this model.

Relative levels of alpha-, beta-, and gamma-mRNA from patients with severe and intermediate beta-thalassemia major.

We have determined the relative quantities of gamma- and beta-mRNAs and the alpha/beta-mRNA ratios in 37 patients with beta-thalassemia major with specific genotypes, namely 8 with a homozygosity for codon (CD) 39 (C-->T), 7 with a homozygosity for IVS-I-110 (G-->A), 5 with a homozygosity for IVS-I-6 (T-->C), for 15 patients with compound heterozygosities for 2 of these 3 mutations, and for 2 patients with the IVS-I-110 (G-->A)/-87 (C-->G) mutations. None had an alpha-thalassemia. Twelve patients had thalassemia intermedia and the remainder, transfusion-dependent severe conditions. Differences in phenotype were observed for compound heterozygotes involving the IVS-I-6 (T-->C) mutation in combination with either the IVS-I-110 (G-->A) or the CD 39 (C-->T) mutations: patients with thalassemia intermedia had a lower alpha/beta-mRNA ratio, about half of that of the patients with severe beta-thalassemia major. This might suggest a higher beta-mRNA synthesis in some patients than in others with the same genotype; mutations in promoter, enhancer, and/or locus control region sequences may be responsible for these differences. In vitro chain synthesis data were too incomplete to be helpful in this study. The RT-PCR procedure allowed the separation of abnormal (extended) mRNA from normal beta-RNA in subjects carrying the IVS-I-110 (G-->T) mutation. The relative quantities of this beta Th-mRNA (% of beta A + beta Th) were determined by scanning of the appropriate autoradiograms; they averaged 25% for homozygotes and about 4% for heterozygotes, indicating a considerable instability of the message.