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H A Büller

Publications and source records attributed to H A Büller.

15 recordsLinked to original sources
PubMed1992

[Pregnancy during residency].

The number of female residents in the Netherlands has steadily increased in recent years. Due to the increased time on waiting lists to enter residency programmes and to the increased duration of training, female residents will be older during their residencies. This will probably result in an increased number of pregnancies during residencies. A questionnaire regarding pregnancy during residency was sent to 191 residents in two university hospitals in the Netherlands. The response rate was 74.3%. Fifty percent of the male and only 19% of the female residents had children. No negative effects of a pregnancy on their training were experienced or anticipated by the residents. However, a negative effect on the functioning of the department was expected. No formal provisions, like replacements were available and many solutions to replace pregnant colleagues depended on the flexibility of the colleagues. The wish to have children was high and equally distributed among male and female residents, 92% and 96%, resp. Given the difficulty to seek a permanent position and to have children after residency, the choice of many female residents will be to have their children during residency. This increase in number of pregnancies requires anticipation of the residency programme directors. They should take the lead in proposing adequate regulations.

Adult↗
PubMed1992

Messenger RNA sorting in enterocytes. Co-localization with encoded proteins.

This study describes the intracellular compartmentalization of three different mRNAs in the polarized rat fetal enterocyte. They encode proteins that are known to be localized within different regions of the epithelial cell namely (i) the apical, membrane-bound glycoprotein, lactase-phlorizin hydrolase (lactase), (ii) the mitochondrially localized enzyme, carbamoylphosphate synthetase (CPS), and (iii) the cytoplasmically localized enzyme, phosphoenolpyruvate carboxykinase (PEPCK). These mRNAs are found in close proximity to their respective protein products, i.e. the apical membrane, mitochondria and cytoplasm, respectively. The significance of these observations is twofold; (i) they indicate that mRNAs are sorted into specific domains of the cytosol of intestinal epithelial cells; and (ii) they imply the presence of two distinct pathways of mRNA targeting one that allows transport of mRNAs that are translated on ribosomes associated with the rough endoplasmic reticulum (lactase mRNA), and the other that allows sorting of mRNAs that are translated on free polysomes (CPS and PEPCK mRNA).

Animals↗
PubMed1992

Lactase gene expression during early development of rat small intestine.

Expression of lactase messenger (m) RNA and protein in rat small intestine during fetal and postnatal development was analyzed using in situ hybridization and immunohistochemistry. Lactase mRNA was first identified at 18 days of development, and lactase protein was first detected at day 20. Lactase mRNA and protein were present along the entire villus. Lactase mRNA increased, reaching a maximum at day 20. Just before birth a decrease in lactase mRNA was observed. In newborn intestine, lactase mRNA was present only from the base of the villus up to the mid-villus region and was undetectable up to the villus tips. Lactase protein continued to be expressed along the entire villus. These data show that expression of lactase mRNA and protein do not parallel, indicating a posttranscriptional control in fetal development. Lactase gene transcription is initiated late in gestation concomitant with villus formation and is exclusively seen in villus epithelial cells. The restriction after birth of lactase mRNA expression to cells at the villus base suggests the occurrence of a previously unknown step in postnatal differentiation of the enterocyte.

Age Factors↗
PubMed1992

Molecular basis of lactase levels in adult humans.

The molecular basis of adult human "lactase deficiency" has long been a subject of controversy. To address this issue, small intestinal biopsies from orienta, black, and white patients were analyzed. Adjacent samples were assayed for lactase and sucrase specific activities and the sucrase/lactase ratio (high ratio signifies lactase deficiency), and the results were compared to lactase steady-state mRNA levels detected in Northern blots probed with a human lactase mDNA. All oriental patients had high ratios and no detectable lactase mRNA. Four black patients had a similar pattern; two with low ratios had detectable mRNA. The group of white patients displayed a range of findings, from high ratio/no mRNA to low ratio/considerable mRNA. Elevated levels of lactase mRNA always correlated with the presence of elevated levels of lactase enzyme activity, suggesting that the difference in levels of adult human intestinal lactase activity among racial groups may be regulated at the level of gene transcription.

Adult↗
PubMed1991

Lactose intolerance and the genetic regulation of intestinal lactase-phlorizin hydrolase.

Lactase-phlorizin hydrolase, which hydrolyzes lactose, the major carbohydrate in milk, plays a critical role in the nutrition of the mammalian neonate. Lactose intolerance in adult humans is common, usually due to low levels of small intestinal lactase. Low lactase levels result from either intestinal injury or (in the majority of the world's adult population) alterations in the genetic expression of lactase. Although the mechanism of decreased lactase levels has been the subject of intensive investigation, no consensus has yet emerged. Recent studies have begun to define the cellular and molecular biology of this enzyme. In animals and humans, a glycosylated precursor is proteolytically cleaved to yield the mature enzyme on the microvillus membrane of the enterocyte, bound to the lipid bilayer only by a hydrophobic anchor sequence. The enzyme hydrolyzes lactose, phlorizin, and glycosylceramides. A decline in lactase specific activity occurs at the time of weaning in most mammalian species; in most humans who have low lactase activity as adults, the decline occurs at approximately 3-5 years of age. In a few human groups, the elevated juvenile level of lactase specific activity persists throughout adulthood. These developmental patterns of lactase expression are most likely regulated at the level of gene transcription.

Animals↗
PubMed1991

Clinical aspects of lactose intolerance in children and adults.

The principal carbohydrate of human milk is the disaccharide lactose. In human and all mammalian species, lactose is hydrolyzed in the small intestine by lactase-phlorizin hydrolase, also abbreviated as lactase. The absence of lactase results in the passage of undigested lactose into the large intestine and is associated with a well-known clinical syndrome: lactose intolerance. Low lactase levels result either from intestinal injury or, as in the majority of world's adult population, from alterations in the genetic expression of lactase. In this review terminology, pathophysiology, symptoms, diagnostic procedures, and therapy of lactose intolerance will be discussed.

Adult↗
PubMed1990

Coordinate expression of lactase-phlorizin hydrolase mRNA and enzyme levels in rat intestine during development.

The development of rat intestinal lactase-specific activity displays a well-known post-weaning decline. In contrast, total lactase activity increases to reach maximal levels around weaning, and remains high subsequently. In order to elucidate the molecular basis for these patterns, a rat lactase cDNA was isolated and characterized, and used in the quantification of lactase mRNA during development. This lactase cDNA uniquely hybridized to a 6.8-kilobase mRNA in the small intestine. To assess the amount of lactase mRNA encoding for lactase enzyme activity in the small intestine, total intestinal RNA was isolated and analyzed by Northern and dot-blot hybridization. The pattern of total lactase mRNA during development followed that of total lactase activity, suggesting that over this time span the level of lactase activity is primarily controlled at the transcriptional level. However, the magnitude of increase of total lactase activity during lactation compared to that of total lactase mRNA suggests that additional mechanisms are involved in regulating lactase levels. Analysis of the regional distribution of lactase mRNA along the small intestine at 14 days revealed that mRNA was high in the proximal three regions, but was dramatically lower in the distal regions. Total lactase activity, in contrast, displayed maximum activity in the mid-intestine with decreased levels both proximally and distally. Thus, lactase activity in the intestine appears to be regulated during development predominantly by transcriptional mechanisms, while alterations during lactation, and along the proximal to distal gradient, are the result of other control mechanisms.

Aging↗
PubMed1990

Glycosylation of lactase-phlorizin hydrolase in rat small intestine during development.

Age-specific changes in glycosylation of rat intestinal lactase-phlorizin hydrolase were analyzed using enzyme immunoprecipitated from microvillus membranes of suckling, weaning, and adult rats, and carbohydrate moieties were examined by lectin affinity binding, metabolic labeling, and neuraminidase treatment. Lectin binding indicated the presence of N-linked and O-linked oligosaccharide chains containing mannose and galactose throughout development. An age-dependent shift in sialic acid and fucose was seen during the period of weaning; no fucose was detectable in lactase-phlorizin hydrolase until after the rats were 20 days of age, whereas sialic acid was reduced in adult lactase-phlorizin hydrolase. The presence of sialic acid in suckling intestines and fucose in adult was confirmed by metabolic labeling with appropriate radioactive precursors. Sodium dodecyl phosphate-polyacrylamide gel electrophoresis analysis of immunoprecipitated lactase-phlorizin hydrolase from the proximal and mid small intestine showed two bands of approximately 220 and 130 kilodaltons in all age groups. In the distal part of the adult small intestine, lactase-phlorizin hydrolase appeared as two bands of similar size to those found in the proximal and mid portions. In contrast, during the suckling and weaning periods, these distal bands were approximately 225 and 135 kilodaltons. [35S]-methionine labeling and fluorography of neonatal intestines confirmed these observations. The size difference between proximal and distal small intestines was virtually eliminated by neuraminidase treatment. These data indicate that the core structure of microvillus membrane lactase-phlorizin hydrolase, consisting of both N-linked and O-linked oligosaccharides, remains constant during development, although terminal sugars shift from predominantly sialic acid during the suckling period to fucose in adulthood. This alteration in glycosylation of the protein occurs in a different pattern from the postweaning decline in lactase specific activity. Consequently, age-dependent changes in glycosylation cannot account for the decrease in lactase-phlorizin hydrolase-specific activity observed during development.

Aging↗
PubMed1990

Lactose intolerance.

Lactose intolerance is a prevalent clinical problem. Low lactase levels result either from intestinal injury, or as in the majority of the world's adult population, from alterations in the genetic expression of lactase-phlorizin hydrolase. Progress is being made in the basic understanding of the molecular and cellular biology of this enzyme and of the scientific basis of clinical syndromes involving low lactase activity.

Animals↗
PubMed1989

Further studies of glycosylation and intracellular transport of lactase-phlorizin hydrolase in rat small intestine.

Previous studies [Büller, Montgomery, Sasak & Grand (1987) J. Biol. Chem. 262, 17206-17211] have demonstrated that lactase-phlorizin hydrolase is inserted into the microvillus membrane (MVM) as a large precursor of approx. 220 kDa, which then undergoes two proteolytic cleavage steps to become the 130 kDa mature MVM protein. In order to assess the role of glycosylation in intracellular transport, the processing of this enzyme has been studied in the presence of castanospermine, an inhibitor of N-linked oligosaccharide modification and subsequent treatment with two endoglycosidases, endo-beta-N-acetyl-glucosaminidase (endo-H) and peptide:N-glycosidase-F (N-glycanase). We now show that the intracellular precursor (205 kDa) undergoes carbohydrate processing (220 kDa) and transport to the MVM where its further proteolytic cleavage is as described. Treatment of the intracellular 205 kDa precursor with either endo-H which cleaves only high-mannose N-linked oligosaccharides, or with N-glycanase, which cleaves both high-mannose and complex N-linked oligosaccharides, results in the conversion of the 205 kDa protein band to one of 195 kDa. These data suggest that the 205 kDa precursor contains only high-mannose N-linked carbohydrates, and that the unglycosylated nascent protein is 195 kDa. In the presence of castanospermine, an intracellular precursor of approx. 210 kDa is observed. When treated with endo-H or N-glycanase, this form also produces a protein of 195 kDa. The transport of the intracellular precursor to the MVM and further proteolytic processing is not blocked by the inhibitor. However, all MVM forms of lactase-phlorizin hydrolase show an increase of approx. 5 kDa. Treatment of these three MVM forms with endo-H indicates the increased presence of high mannose oligosaccharides in comparison with non-castanospermine-treated forms. The susceptibility to endo-H of the 130 kDa MVM band synthesized in the absence of castanospermine implies the presence of high-mannose N-linked oligosaccharides in the mature form of lactase-phlorizin hydrolase. Incubation of these MVM forms with N-glycanase further reduces their electrophoretic mobility, indicating the presence of complex N-linked oligosaccharides in the MVM forms, in contrast with the intracellular precursor. Altered glycosylation reduces but does not abolish intracellular transport of lactase-phlorizin hydrolase to the MVM.

Alkaloids↗
PubMed1989

New insights into lactase and glycosylceramidase activities of rat lactase-phlorizin hydrolase.

Lactase-phlorizin hydrolase, a small intestinal disaccharidase, has been considered mainly an enzyme important only for the hydrolysis of lactose. After weaning in most mammals lactase-specific activity falls markedly, and, functionally, adult mammals are considered to be lactase deficient. However, the persistence of low levels of lactase activity in adulthood has never been explained. In addition, it has been suggested that lactase-phlorizin hydrolase is associated with glycosylceramidase activity when the enzyme is prepared by column chromatography, but it is unclear whether this represents copurified activities or two catalytic sites on one peptide. The developmental patterns of lactase-phlorizin hydrolase and other disaccharidases were investigated in homogenates of total rat small intestine; lactase and several glycosylceramidases were measured in immunoprecipitates from these homogenates using a monoclonal antibody. The developmental pattern of total lactase activity showed a steady 2.3-fold increase to adult levels (specific activity decreased eightfold), whereas total phlorizin-hydrolase activity increased 10.7-fold (specific activity decreased threefold). As expected, levels of both total and specific sucrase and maltase activities increased during development. In lactating rats total lactase activity showed a significant increase compared with adult males. The developmental pattern of the enzyme activities for the glycolipid substrates was similar to that found for lactase, and the immunoprecipitated enzyme showed a 40- to 55-fold higher affinity for the glycolipids than for lactose. Galactosyl- and lactosylceramide inhibited lactose hydrolysis by 38%, without a competitive pattern, suggesting two different active sites for lactose and glycolipid hydrolysis, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗
PubMed1989

Suckling rat colon synthesizes and processes active lactase-phlorizin hydrolase immunologically identical to that from jejunum.

To identify potential tissue-specific characteristics of intestinal glycoprotein synthesis and processing, rat intestinal lactase-phlorizin hydrolase (L-Ph) was studied after pulse-labeling of colonic explants from 5-d-old suckling rats in organ culture and the data compared to similar studies in rat jejunum. Histologic sections of 5-d-old proximal colon showed villus-like structures lined with columnar epithelial cells. Lactase and phlorizin hydrolase activities showed tissue-specific developmental patterns. Using a MAb to small intestinal L-Ph, we were able to immunoprecipitate from colon at different ages a protein that hydrolyzed lactose and phlorizin, and whose activity was not inhibited by p-chloromercuribenzoate. After pulse-labeling for 60 min and chase for 30 min, immunoprecipitated L-Ph from total homogenates of rat colonic explants appeared on fluorography of SDS-PAGE as one band of approximately 205 kD. With increasing time of chase, it took 240 min before the precursor form was converted to the intermediate form (equivalent to the 180-kD form in jejunum) and the mature form (equivalent to the 130-kD form in jejunum), although these conversions in the jejunum were observed within 60 min of chase, and only 30 min of pulse labeling. When compared on SDS-PAGE to immunoprecipitated jejunal L-Ph, the precursor form in the colon had a slightly higher apparent mol wt than the corresponding precursor form found in the endoplasmic reticulum-Golgi fraction of the jejunum. The intermediate as well as the mature L-Ph forms in the colon were also both somewhat higher in apparent molecular weight than the same bands in the microvillus membrane fraction from jejunal explants. Removal of N-linked oligosaccharides from jejunum and colonic forms of L-Ph produced bands on SDS-PAGE with identical mobility, suggesting that the proteins were the same. The data demonstrate that, in neonatal colon, enzymatically active L-Ph undergoes biosynthetic and processing events similar to those in the jejunum. During early life, colonic L-Ph may function in the salvage of lactose not absorbed in the small intestine.

Animals↗
PubMed1987

Biosynthesis, glycosylation, and intracellular transport of intestinal lactase-phlorizin hydrolase in rat.

The biosynthesis of rat intestinal lactase-phlorizin hydrolase was studied by pulse-labeling of jejunal explants from 5-day-old suckling rats in organ culture. Explants were either continuously labeled with [35S] methionine for 15, 30, and 60 min or pulse-labeled for 30 min and chased for various periods of time up to 6 h in the presence or absence of protease inhibitors (PI), leupeptin, phenylmethylsulfonyl fluoride, and soybean trypsin inhibitor. Lactase-phlorizin hydrolase was immunoprecipitated from microvillus membrane (MVM) and ER-Golgi fractions with monoclonal antibodies. After pulse-labeling, lactase-phlorizin hydrolase from the ER-Golgi fraction appeared on SDS-PAGE as one band of approximately 220 kDa, regardless of the presence or absence of PI in the culture media. The 220-kDa protein band could also be labeled after incubation with [2-3H]mannose. In the absence of PI, the 220-kDa band appeared in the MVM by 30 min chase, simultaneously with a 180-kDa band, and by 60 min of chase an additional band of 130 kDa was seen. With increasing time of chase, the relative intensity of the 130-kDa band increased, whereas that of the 220-kDa band decreased, suggesting a precursor-product relationship. When PI were added to the medium, the formation of the 180-kDa band was not affected, but the conversion of the 180-kDa protein to the 130-kDa protein was virtually blocked. These findings suggest that lactase-phlorizin hydrolase is initially synthesized as a glycosylated precursor of 220 kDa, which is transported to the MVM. There it undergoes the following two cleavages: first, to the 180-kDa form, which is not prevented by PI used in these experiments, and second, to the 130-kDa form inhibited by PI.

Animals↗