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C Y Cheng

Publications and source records attributed to C Y Cheng.

At least 181 records · Page 10Linked to original sources
PubMed1986

Rat testicular testibumin is a protein responsive to follicle stimulating hormone and testosterone that shares immunodeterminants with albumin.

During a search for hormonally responsive products in media from Sertoli cell enriched cultures, a major follicle stimulating hormone responsive and testosterone-responsive protein was identified and designated CMB-1. The results of the present study indicate that this protein is related immunologically to rat albumin and rat alpha-fetoprotein (AFP) and is concentrated in the testis of the adult rat. CMB-1 was therefore termed testibumin. Testibumin was purified from Sertoli cell enriched cultures to apparent homogeneity by sequential high-performance liquid chromatography on anion-exchange, chromatofocusing, gel permeation, and hydroxylapatite columns. The purified protein consists of two concanavalin A (Con A) reactive forms: one which does not interact with Con A and the other which binds to this lectin and is eluted with methyl alpha-mannoside. Testibumin is a monomer with an apparent molecular weight of 69,000 and a pI ranging between 4.5 and 4.85. The heterogeneity of this protein was further demonstrated by crossed-immunoelectrophoresis and two-dimensional gel electrophoresis. A monospecific antiserum and highly purified testibumin were used to develop a specific radioimmunoassay which permitted studies of the hormonal responsiveness of Sertoli cell enriched culture and of the content of testibumin in the reproductive tract fluids in vivo. Even though testibumin was found in serum of both sexes, it was highly concentrated in the testicular and epididymal compartments in adult rats. This protein was compared to rat serum albumin and rat AFP immunologically. With the use of immunoblots, antiserum developed against testibumin showed partial cross-reactivity with albumin and AFP when these latter proteins were denatured and were present in amounts several orders of magnitude greater than testibumin. The extent of this cross-reactivity was then examined by comparing the ability of native and S-carboxymethylated albumin to compete with 125I-testibumin for binding to a monospecific testibumin antiserum. It was shown that the unfolded derivative of albumin showed partial cross-reactivity with testibumin. We conclude that testibumin is immunologically related to albumin and AFP as these latter proteins are related to one another and that testibumin is possibly the homologue of albumin in the seminiferous tubular compartment.

Animals↗
PubMed1986

Identification of hormonally responsive proteins in primary Sertoli cell culture medium by anion-exchange high performance liquid chromatography.

Twenty day-old rats were used to prepare Sertoli cell-enriched cultures which were grown for 4 days in serum-free medium supplemented with human transferrin, epidermal growth factor, and insulin. Proteins in the medium were first fractionated by anion-exchange HPLC. Forty-five to 50 fractions were collected, and each was examined by electrophoresis in sodium dodecyl sulfate-containing polyacrylamide gels. Using silver nitrate as a stain, more than 30 proteins were routinely identified. Rat androgen binding protein and rat transferrin were measured by immunoassays. When the cultures were supplemented with either testosterone, FSH, or both hormones and the media fractionated by HPLC and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, these proteins could be tentatively categorized as follows: unresponsive to added hormones; responsive primarily to testosterone; responsive primarily to FSH; or responsive to testosterone or FSH but maximally responsive to both hormones. This report provides a method for identifying proteins in Sertoli cell-enriched cultures and has allowed us to show for the first time that many are responsive to hormones. The procedure takes advantage of the large loading capacity of the HPLC column to allow fractionation of liter quantities of medium. The high resolving power of the column permits preparation of fractions which are highly enriched with one or a limited number of proteins. The purification of many of these proteins is now feasible. The approach described is generally applicable for isolation of proteins from media from a variety of cells.

Androgen-Binding Protein↗
PubMed1986

Measurement of a follicle-stimulating hormone-responsive protein of Sertoli cell origin using an enzyme-linked immunoblot assay.

Many proteins secreted by Sertoli cell-enriched cultures are maximally stimulated by a combination of FSH and testosterone. Since very few are stimulated primarily by FSH, we thought it pertinent to identify such proteins. Sertoli cell-enriched cultures were prepared from testes of 20-day-old rats and grown in serum-free medium containing insulin, transferrin, and epidermal growth factor and in such medium supplemented with FSH, testosterone, or FSH plus testosterone. Media were fractionated using HPLC, and proteins were identified by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. A protein designated CMB-2, with an apparent mol wt of 22,000, was shown to increase in response to FSH. Antiserum was raised using denatured protein eluted from SDS-polyacrylamide gels as the antigen, and a specific immunoassay using a combination of SDS-polyacrylamide gel electrophoresis and Western blotting was developed. The production of CMB-2 by primary Sertoli cell-enriched cultures was found to increase in a dose-dependent manner in response to FSH (30-1000 ng/ml); secretion was not significantly affected by testosterone (2 X 10(-13) M). An investigation of the tissue distribution of CMB-2 showed that the puberty, CMB-2 is secreted into the rete testis and accumulates in the epididymis in high concentration. We conclude that CMB-2 will be a useful marker to study the action of FSH on the rat testis.

Androgen-Binding Protein↗
PubMed1985

There are two forms of androgen binding protein in human testes. Comparison of their protomeric variants with serum testosterone-estradiol binding globulin.

To determine how the androgen binding protein in human testes (hABP) is related to the serum protein, testosterone-estradiol binding globulin (hTeBG), both proteins were isolated and compared. The hABP in extracts of human testes was composed of two molecular species based on concanavalin A (ConA)-Sepharose chromatography. Form I hABP did not interact with ConA while Form II hABP bound to ConA and eluted with alpha-methylmannoside. Form I and Form II hABP from five batches of testes were then purified approximately 30,500- and 30,000-fold to apparent homogeneity by high-performance liquid chromatography and compared with hTeBG isolated from human pregnancy serum. Fractionation of both forms of hABP and hTeBG by polyacrylamide gel electrophoresis in the absence of sodium dodecyl sulfate suggested that the native forms of these proteins were indistinguishable. However, analysis of the purified proteins on sodium dodecyl sulfate-containing polyacrylamide gels indicated that all three were dimers and that each was composed of monomers of at least two sizes which were not present in equimolar concentrations. Two distinctive monomers or protomers of each protein were designated as heavy (H) and light (L) according to their electrophoretic mobilities on sodium dodecyl sulfate-polyacrylamide gels. The H and L protomers of Form I hABP showed apparent molecular weights of 55,000 and 52,000, respectively, in all preparations and were usually present in a 4:5 ratio (H:L). The two components of Form II hABP had apparent molecular weights of 53,000 and 48,000, respectively, and existed in a ratio of approximately 20:1. These two components could not be distinguished in some preparations where Form II hABP migrated as a broad band rather than as distinct protomers. By contrast, hTeBG, which was similar to Form II hABP with respect to ConA binding, always exhibited discrete H and L protomers in a 10:1 ratio. Photolysis of these highly purified proteins with delta 6-[3H]testosterone resulted in specific covalent labeling of their binding sites, confirming that the products identified by silver staining and immunoblotting were indeed steroid binding proteins. The H and L protomers of Form I hABP and hTeBG were separated and examined by peptide mapping using Staphylococcus aureus protease V8 and chymotrypsin. The comparison of the respective fragmentation patterns of protomers indicated that Form I hABP and hTeBG contained distinctive peptides.(ABSTRACT TRUNCATED AT 400 WORDS)

Androgen-Binding Protein↗
PubMed1985

The role of the carbohydrate moiety on the size heterogeneity and immunologic determinants of human testosterone-estradiol-binding globulin.

Human testosterone-estradiol-binding globulin (hTeBG) has been purified to apparent homogeneity by several laboratories using procedures which, in most instances, were labor intensive. In this report, hTeBG was purified from pregnancy serum by a newly developed two step procedure involving sequential affinity chromatography and ion-exchange high performance liquid chromatography (ion-exchange HPLC). The purity of the final product was confirmed by silver stained SDS-polyacrylamide gel and reverse phase HPLC monitored at 206 nm. hTeBG purified by ion-exchange-HPLC maintained binding activity by Dextran coated charcoal (DCC) assay and size heterogeneity on SDS-polyacrylamide gels which were indistinguishable from those of the proteins purified by conventional chromatography. Removal of the carbohydrate moiety from the molecule by both enzymatic and chemical treatment reduced the apparent molecular size and eliminated lectin binding of hTeBG subunits. Deglycosylation did not, however, abolish or alter the distribution of the protomeric forms of this subunit. We conclude that hTeBG is a dimer whose monomer exhibits two protomeric forms which is not a result of carbohydrate heterogeneity. In addition, disialylated and deglycosylated hTeBG exhibited antigenic determinants identical to the native protein.

Carbohydrates↗
PubMed1985

Purification of testosterone-oestradiol-binding globulins from mammalian sera by anion-exchange high performance liquid chromatography.

Testosterone-oestradiol-binding globulin (TeBG) has been isolated from serum or plasma of several species using procedures that yielded highly purified protein, but which required multiple and tedious chromatographic steps. In this report we describe a procedure for the isolation of TeBG which involves two chromatographic steps: androgen affinity chromatography followed by anion-exchange high performance liquid chromatography (anion-exchange HPLC). The purity of the final product was confirmed by silver staining following fractionation on sodium dodecyl sulphate-containing polyacrylamide gels. The size heterogeneity and specific binding activity of TeBGs purified from human, rabbit, or bull serum (or plasma) by this technique was indistinguishable from preparations obtained by conventional chromatography. The present technique shortened the entire purification procedure to about 5 working days and yielded milligram quantities of highly purified protein. Bases on our experience with serum or plasma from the human, rabbit, and bull, this approach should be suitable for isolation of TeBG from a wide range of species.

Animals↗
PubMed1985

Sex hormone-binding globulin changes during the menstrual cycle.

Although sex hormone-binding globulin (SHBG) production is stimulated by estrogen, no change in SHBG has been demonstrated during the menstrual cycle. To further study possible cyclic changes in serum SHBG, 12 women with a normal menstrual and fertility history had daily SHBG measurements during a menstrual cycle. SHBG was measured by dextran-coated charcoal saturation analysis and RIA. Serum LH was measured by mouse Leydig cell bioassay and RIA, and FSH, estradiol (E2), and progesterone were determined by RIA. In 10 women, a significant increase in mean SHBG by both methods occurred during the luteal phase of the cycle, immediately after the preovulatory increase in serum E2 (P less than 0.001). Two women had no SHBG increase; although each had a significant rise in serum E2 before the LH surge, luteal phase E2 levels were similar to those in the early follicular phase. In one of these women, a rise in SHBG was demonstrated by RIA. This study demonstrates that SHBG changes during the menstrual cycle in association with E2 changes, and it appears to be a marker for the endogenous estrogen changes that occur in normal ovulating women.

Adult↗
PubMed1984

The heterogeneity of rat androgen-binding protein in serum differs from that in testis and epididymis.

Fractionation of testicular, epididymal, and serum extracts containing rat androgen-binding protein (rABP) on a Concanavalin A-Sepharose (Con A) column resolved two peaks of immunoreactive protein. The first peak was present in the void volume, and the other was bound by the column and specifically eluted by alpha-methyl-D-glucoside. These two peaks of immunoreactive rABP have been designated form I and form II for the portions of rABP that do not and do bind, respectively, to Concanavalin A. In the course of studying this heterogeneity, we observed that the distribution of the two forms of rABP was the same in the blood and cytosols prepared from testis and epididymis of young rats before the formation of the blood-testis barrier; that is, the ratio of form I to form II ranged from 1:1 to 1:2. Similar heterogeneity was observed in extracts of the reproductive tract from mature animals. However, the blood of adult rats contained reduced amounts of form I relative to form II, so that their ratio was about 1:5. Subsequent studies of infertile rats heterozygous for the Hre gene (Hre/ +), in which total rABP secretion was decreased, and of their normal littermates, indicated that the reduced amount of form I ABP in the sera of mature rats is typical of adult animals regardless of strain or genetic abnormality. The reduced amount of form I relative to form II observed in the blood of adult rats could result from either reduced secretion or increased metabolic clearance of form I in the blood compartment. To distinguish between these possibilities, the blood clearance of the two forms was estimated after orchiectomy. The disappearance rate of form I was not significantly different from that of either form II or unfractionated serum. These results are consistent with reduced release into blood of form I relative to form II rABP rather than increased clearance of form I in adult animals.

Aging↗
PubMed1984

Human testicular androgen-binding protein shares immunodeterminants with serum testosterone-estradiol-binding globulin.

In the human, there are two glycoproteins, testosterone-estradiol-binding globulin (hTeBG) and androgen-binding protein (hABP), which bind testosterone. Although these two proteins have similar physicochemical properties, they can be distinguished on the basis of origin and lectin binding. hTeBG is made in the liver and exhibits high affinity for Concanavalin A (Con A), while hABP from the testes is only partially bound to this lectin. That is, when testicular extracts were applied to Con A-Sepharose columns, a portion of the testosterone-binding material showed no interaction with the lectin and eluted in the void volume (peak I), while the remainder interacted strongly and could be eluted with alpha-methyl-D-glucoside (peak II). These observations are consistent with the proposal that peak I contains only hABP, whereas peak II contains hTeBG and/or hABP with carbohydrate units that permit binding to Con A. To further study the properties of these binding proteins, a hTeBG RIA using a monospecific antiserum was employed to compare the proteins in testes and serum. The results indicated that the testosterone-binding activities in peaks I and II of testicular extracts could not be distinguished immunologically from hTeBG in sera of normal women. These findings suggested that hTeBG and hABP share common epitopes. We next determined whether hABP was secreted into the blood or amniotic fluid by fractionating these fluids in Con A-Sepharose columns. Unlike testicular extracts, male serum and amniotic fluid contained single immunoreactive and steroid-binding species which bound specifically to Con A. We conclude from these observations that hABP (peak I), peak II activity, and hTeBG have common immunodeterminants and that if hABP is secreted into the blood of men, then its carbohydrate chains bind to Con A, making it indistinguishable from hTeBG under these conditions.

Adult↗
PubMed1983

Demonstration of heavy and light protomers of human testosterone-estradiol-binding globulin.

Human testosterone-estradiol-binding globulin (hTeBG) was purified from pregnancy serum by sequential ammonium sulfate precipitation, affinity chromatography, and hydroxylapatite chromatography. An overall purification of 2800-fold was achieved with a 27% total yield. Apparent homogeneity of the final product was shown by polyacrylamide gel electrophoresis with or without sodium dodecyl sulfate (SDS). The equilibrium dissociation constant (Kd) at 4 degrees C for 5 alpha-dihydrotestosterone (DHT) was estimated to be 1.94 +/- 0.95 X 10(-9) M. Analysis of the purified protein revealed microheterogeneity with regard to size on polyacrylamide gel in the presence of SDS and to charge on isoelectric focusing gels. The apparent molecular weight of native hTeBG determined by gradient gel electrophoresis was 115,000. SDS-polyacrylamide gel electrophoresis indicated that hTeBG is comprised of two molecular weight components of 53,000 and 46,000, which are designated as heavy (hTeBGH) and light (hTeBGL) protomers, respectively. Photolysis of purified hTeBG with [1,2-3H]17 beta-hydroxy-4,6-androstadien-3-one [( 3H]delta 6-testosterone) resulted in specific labeling of its binding sites. Analysis of photolabeled products by SDS-polyacrylamide gel electrophoresis revealed two radioactive products with electrophoretic mobilities identical to those of the hTeBGH and hTeBGL. The ratio of hTeBGH to hTeBGL was about 10:1. The H and the L protomers were separated and examined by peptide mapping using protease V8 and chymotrypsin. Comparison of the fragmentation patterns produced by these proteases revealed that hTeBGH and hTeBGL components were nearly identical. Removal of sialic acid or carbohydrate residues from hTeBG did not affect the presence of two molecular components. Isoelectric focusing of native hTeBG demonstrated three isoelectric variants with pIs at 4.75, 4.85 and 4.90. After treatment with neuraminidase and other glycosidases, only two isoelectric species were observed with more alkaline pIs. Although purified hTeBG appeared heterogeneous with regard to size and charge, it was remarkably homogeneous in its ability to absorb to Concanavalin A-Sepharose. We conclude that hTeBg, like the androgen binding proteins of the rabbit and rat, is a dimer whose monomer exhibits two protomeric forms.

Chemical Phenomena↗
PubMed1983

Acute fatty liver of pregnancy--survival despite associated severe preeclampsia, coma and coagulopathy.

Acute fatty liver of pregnancy is the least common and most serious cause of jaundice of pregnancy. It is rare, fewer than 100 cases having been reported (Koff, 1981). There is a high probability that it is frequently misdiagnosed and categorized vaguely as some form of variant of 'toxaemia of pregnancy' which is not surprising because of its late-pregnancy onset and the production of multi-system manifestations (Holzback, 1976). In the case reported here, there was associated severe preeclampsia with fetal intrauterine death but maternal survival.

Adult↗
PubMed1983

Radioimmunoassay of testosterone-estradiol-binding globulin in humans: a reassessment of normal values.

Antiserum against human testosterone-estradiol-binding globulin (hTeBG) was prepared in rabbits, and its specificity was demonstrated by crossed immunoelectrophoresis. A RIA for the measurement of hTeBG in serum was developed. With this assay, hTeBG was readily measured in 1-2 microliters serum. Interassay coefficients of variation for pools of serum from men, women, and women in late pregnancy were 7.7%, 5.5%, and 6.1%, respectively. Interassay coefficients of variation for the same pools were 10%, 12%, and 12%, respectively. The TeBG levels in a number of nonselected subjected determined by the present method show good correlations with those obtained by steady state polyacrylamide gel electrophoresis and dextran-coated charcoal assay. The concentrations of TeBG determined by the RIA in sera from men, women, and women in late pregnancy were 18 +/- 9 (n = 12), 54 +/- 13 (n = 8), and 374 +/- 55 (n = 6) pmol/ml (mean +/- SD), respectively.

Amniotic Fluid↗
PubMed1983

The hormonal and cellular control of Sertoli cell secretion.

Sertoli cells synthesize and secrete a number of cell-specific products including androgen binding protein (ABP), as well as "serum proteins" such as transferrin. The secretion of these proteins is regulated by extra-testicular hormones such as FSH and insulin; Leydig cell-produced steroids and proopiomelano-cortin-derived peptides; and the presence of peritubular myoid cells and/or factors secreted by these cells. Many of the Sertoli cell proteins are secreted in a cyclic fashion during the different stages of the spermatogenic cycle suggesting communication between Sertoli cells and developing germ cells. The availability of quantitative measurements for Sertoli cell-specific proteins such as ABP make it feasible to follow Sertoli cell function in vivo by measuring these products in serum. A bidirectional secretion of proteins by Sertoli cells is proposed to explain the presence of specific peptides in the male reproductive tract and blood.

Androgen-Binding Protein↗
PubMed1982

Partial purification of components from fasting human blood serum which stimulate the forward migration of human spermatozoa.

Components from fasting human serum that could stimulate the forward migration of human spermatozoa were isolated by gel filtration on Sephadex G-25. The results suggest that these components may be used to enhance sperm forward migration and, hence, pregnancy rate in artificial insemination with husband's semen. (AIH) especially in cases where the sperm forward migration is not optimal.

Blood Physiological Phenomena↗
PubMed1982

Quantitative structure-activity relationships of aromatic esters of 1-methyl-4-piperidinol as analgesics.

Substituted benzoic acid esters of 1-methyl-4-piperidinol showed analgesic activity when assayed by the mouse hot-plate methods, the more potent ones falling in the morphine-codeine range. To understand how substituents on the aromatic ring affect the analgesic potency, quantitative structure-activity correlations were carried out on a series of 44 derivatives. Among the various substituent parameters included in the study, Lortho (Length of ortho-substituents) and B1 (minimal width of substituents) or E8 at meta and para positions gave negative correlation with the potency, while lipophilicity (especially pi meta) and the ability of being a hydrogen-bond acceptor enhanced the potency. Based on the QSAR results, a substitution pattern of the phenyl group was defined for optimal activity. Implications on drug-receptor interactions and the possible binding mode of these compounds were discussed.

Analgesics↗