STUDY ON REPRODUCTIVE ENDOCRINOLOGY OF HUMAN PLACENTA--CULTURE OF HIGHLY PURIFIED CYTOTROPHOBLAST CELL IN SERUM-FREE HORMONE SUPPLEMENTED MEDIUM

A new method of long-term culture of cytotrophoblast cells in serum-free medium has been developed. Cytotrophoblast cells were isolated with cold trypsin and purified by unit gravity sedimentation through BSA density gradients. The cells were cultured in the FD medium with supplement of EGF, insulin, transferrin and sodium selenite. They could survive over three weeks. The results showed that both EGF and insulin stimulated hCG and progesterone secretion and that sodium selenite elevated hCG output but not progesterone secretion. Transferrin produced synergistic effect with EGF and insulin on hCG and progesterone secretion but it was ineffective when used alone. This study demonstrates that the four growth factors mentioned above are essential for the survival of cytotrophoblast cells in vitro. It is therefore suggested that EGF, insulin and selenium may possibly be involved in the regulation of hCG and progesterone secretion in the human placenta.     

PARADOXICAL EFFECT OF A GnRH AGONIST ON STEROIDOGENESIS IN CULTURED MONKEY GRANULOSA CELLS

In this study we demonstrate: (i) The GnRH agonist exerts a direct dose-dependet stimulative effect on the aromatase activity and progesterone production in cultured monkey granulosa cells; (ii)the stimulative effect on steroidogenesis can be completely blocked by concomitant treatment with a GnRH antagonist, suggesting that the actions of GnRH are mediated through stringent stereospecific recongnition sites; (iii) in addition to the stimulative effect, the GnRH agonist in the presence of gonadotropins also exerts an inhibitory effect, even though the peptide by itself is more effective in the stimulation of steroidogenesis, and the stimulation of gonadotropin on steroidogenesis could be gradually restored by decreasing the concentration of the GnRH agonist in the culture; and (iv) paradoxical effect can also be observed in the presence of cAMP-inducing agents, suggesting that the inhibitory action of the peptide on gonadotropin-induced steroidogenesis is localized at a step distal to the stringent reco     

THE BIOLOGICAL FUNCTION OF RABBIT BLASTOCYST PEPTIDES (RBPs)

The inhibitory effect of rabbit blastocystic peptides (RBPs) on lymphocyte transfor-mation was studied by the method of measuring the incorporation of ~3H- thymidine intoDNA. The results indicated that RBPs inhibited PHA-stimulated rat and human lympho-cyte transformation in vitro. In the concentration ranging from 40 to 200 μg/2.5 ml, theinhibition was dose-dependent. No obvious inhibitory action was found with hCG (16- 128IU/2.5 ml), progesterone (250 ng- 1 μg/2.5 ml) and pregnant rabbit serum. It was furtherdemonstrated that RBPs at a higher dosage (200 μg/ml) was inhibitory to PGF_(2α) secretion.On the other hand, the ~3H- leucine incorporation of rabbit endometrium was enhanced bythese peptides, and this action could be blocked by the addition of actidine.     

PHARMACOLOGICAL STUDY OF AN IRREVERSIBLE PARTIAL AGONIST OF OPIATE RECEPTORS, A-α-CAO

A-α-CAO induces weak analgesia with very short duration in mice and is able to antagonize the analgesic effect of morphine (Mor) up to 3—4 days after a single injection. No tendency of dependence has been observed. It acts as a partial agonist on MVD with Ke value of 9×10~(-9) mol/L. Its antagonist effect remains after several washes and its agonist effect cannot be reversed by naloxone (Nx), provided the incubation time or the concentration of the agent is sufficient. On isolated GPI, A-α-CAO is a pure agonist with IC_(50) of 5.7×10~(-10) mol/L; this agonist effect cannot be removed by washing but can be reversed by Nx. On RVD and RbVD, it has antagonist effect against β-endorphine (β-end) and US0488H, which cannot be washed out easily, and the pA_2are 7.5 and 7.6 respectively. A-α-CAO also inhibits the specific binding of ~3H-etorphine (~3H-Etor) to the P_2 fraction of the mouse brain membrane with an IC_(30) of 3.2×10~(-9) mol/L. The inhibition on the high affinity binding sites of ~3H-Etor     

EFFECTS OF CHANGES IN PATTERN OF LHRH PULSE ON LH SECRETION BY PERFUSED ANTERIOR PITUITARY CELLS OF MALE RATS

In the present study, a perfusion system of dispersed cells was used to investigate theeffects of LHRH pulse amplitude and frequency, and LHRH continuous stimulation on LHsecretion by anterior pituitary cells of adult male rats. The results have shown that, in therange of LHRH concentrations from 1×10~(-10) to 1 ×10~(-6) mol/L, the dose-response curve of LHsecretion was linear. LHRH pulse frequency generated a biphasic LH response: increasingLHRH pulse frequency increased the basal LH secretion and decreased LH/pulse. When1 ×10~(-9), mol/L or greater LHRH was given at frequencies of 3 pulses/h or higher, it wasobserved that a maximal LH peak was induced and then the LH release declined progres-sively to its LH basal level, i.e. LHRH self-priming effect and LH desensitization occurred.Enhancement of amplitude of LHRH pulses could reduce pulse frequency required for prim-ing. Increases in frequency of LHRH pulses with high amplitude would provoke the prim-ing effect more quickly. In addition, continuou     

STUDY ON REPRODUCTIVE ENDOCRINOLOGY OF HUMAN PLACENTA (Ⅱ)——HORMONE SECRETING ACTIVITY OF CYTO-TROPHOBLAST CELLS

The capability of cytotrophoblast cells to produce hCG, progesterone, estrogen, cGnRH and β-endorphin in vitro has been demonstrated in serum-free culture medium. Before experiment, a 24-h preculture was carried out in order to remove the endogenous hormones of the tissue. During a period of 8 days' culture, the cytotrophoblast cells could constantly produce a small amount of hCG. The production of progesterone rose rapidly and became doubled within six days. The estrogen secretion showed a similar pattern in the presence of androstenedione, a precursor of estrogen, indicating the elevation of aromatase activity in the cells. The elevation of the enzyme activity has been further demonstrated not to be induced by androstenedione. In both cytotrophoblast and syncytiotrophoblast cell cultures, cGnRH was only detected in the culture of cytotrophoblast cells, with a value up to 4 pg/105 cells/24 h. However, β-endorphin was identified both in cytotrophoblast and syncytiotrophoblast cells. Its content increa     

COULOMETRIC TITRATION OF MYLIS

A method for the determination of Mylis by coulometrictitration was developed.The titration was carried out in a mixture of1 mol/L KBr-2 mol/L HCl(1:1)and bromine was generated at the anode,Mylis reacted with one mole of bromine with a n value of 2.The advantages of this method are simple and rapid,it can be used toanalyze small amount of sample.     

CRITICAL CONCENTRATIONS OF α(1→3)-D-GLUCAN FROM LENTINUS EDODES IN NaOH AQUEOUS SOLUTION

Critical concentrations of α-(1→3)-D-glucan L-FV-Ⅱ from Lentinus edodes were studied by viscometry andfluorescence probe techniques. The dependence of the reduced viscosity on concentration of the glucan in 0.5 mol/L NaOHaqueous solutions with or without urea showed two turning points corresponding to the dynamic contact concentration c_s andthe overlap concentration c~* of the polymer. The values of c_s and c~* were found to be 1×10~(-3) g cm~(-3) and 1.1×10~(-2) g cm~(-3),respectively, for L-FV-Ⅱ in 0.5 mol/L NaOH aqueous solutions. The two critical concentrations of L-FV-Ⅱ in 0.5 mol/LNaOH aqueous solutions were also found to be 1.2×10~(-3) g cm~(-3) fbr c_s and 9.2×10~(-3) g cm~(-3) for c~* from the concentrationdependence of phenanthrene fluorescence intensities. The overlap concentration c~* of L-FV-Ⅱ in 0.5 mol/L NaOH aqueoussolutions was lower than that of polystyrene with same molecular weight in benzene, owing to the fact that polysaccharidetends to undergo aggregation caused by intermolecular hydrogen bonding. A normal viscosity behavior of L-FV-Ⅱ in 0.5 mol/L urea/0.5 mol/L NaOH aqueous solutions can still be observed in an extremely low concentration range at 25℃.     

FLUORIMETRIC DETERMINATION OF TETRACYCLINE IN SERUM AND URINE

The Eu-Tetracycline(TC)-TOPO-sodium dodecyl sulfonate(SDS)system wasstudied,Experiments showed that the maximum fluorescence intensity was obtainedin the pH range of 7.5-8.7 and the concentrations of Eu~(a+),TOPO and SDS are1.0×10~(-6)mol/L,1.0×10~(-8)mol/L and 1.0×10~(-8)mol/L,respectively,This fluorescencesystem can be used for the determination of TC in serum and urine,Beer's Law isobeyed in the range of 2.0×10~(-8)mol/L=1.0×10~(-6)mol/L for the concentration of TC.The determination limit is 1.2×10~(-8)mol/L,The composition and the luminescencemechanism were discussed.     

INHIBITIVE ACTION OF INORGANIC COMPOUND ON CORROSION OF ALUMINIUM IN HYDROCHLORIC ACID

The inhibitive action of CaSO_4 towards corrosion of aluminium in 1 mol/L HCl hasbeen inveetigated by using fluorometrio and weight loss techniques.The results show thatthe inhibitor exhibits an outstanding function of inhibition,especially at higher tempe-ratures,and it was found to be predomiantly of cathodic rate control.Ths mechanism ofinhibition and the effect of CdSO_4 on the corrosion kinetics are also discussed.     

STUDY ON THE MECHANISM OF INTERFERON ACTION (Ⅺ)——EFFECT OF pppA2''''p5''''A2''''p5''''A ON THE LEVEL OF cAMP AND cGMP IN MACROPHAGES

In this paper, the increase of cellular cAMP and cGMP levels in macrophages induced bypppA2'p5'A2'p5'A (briefly 2'-5'P_3A_3) is first reported. The optimal concentration of 2'-5'P_3A_3 for the elevation of cellular cGMP to the highest level is 10~(-7)-10~(-6)mol/L, while thatfor cAMP is 10~(-7)mol/L. The time for cGMP to reach its peak value is 15 min and that forcAMP is 2 h, when the cells are treated with 2'-5' P_3A_3 at 10~(-7)mol/L, which is the optimalconcentration for developing biological effect of macrophages (phagocytosis). These resultssuggest that cGMP and cAMP may be related to, or may be the mediators for, 2'-5'P_3A_3action.     

STUDY ON INDUCTION OF DIFFERENTIATION OF A HUMAN MEGAKARYOBLASTIC LEUKEMIA CELL LINE (HIMeg) BY 1,25-DIHYDROXYVITAMIN D_3

It is reported that 1,25-dihydroxyvitamin D_3(1,25(OH)_2D_3), a physiological factor, has aninductive effect on the differentiation of a novel human megakaryoblastic leukemia cell line(HIMeg) in vitro. At the concentrations ranging from 10~(-9) to 10~(-6) mol/L, 1,25(OH)_2D_3 showedinhibition of proliferation on HIMeg cells which was demonstrated by count of survivalcells and cloning efficiency. Meanwhile, using light/electron microscopy, stain of cytochem-istry (including immunoenzymatic technique) and flow cytometry, we found that HIMeg cellscould be further induced into more mature cells in megakaryocytic lineage confirmed by aseries of evidence, including the changes of cell morphology/structure and cytochemistry,increased expression of differentiation antigens on the cell surface, and polyploidization.So, it is possible for 1,25(OH)_2D_3 to promote the differentiation of the cells in megakaryo-cytic lineage in vivo and to be used to treat acute megakaryoblastic leukemia and other di-seases with mal     

Electrochemical Behavior of Mitoxantrone and Its Determination at a Pt／C Implanted Modified Microelectrode

In a 0.02 mol/L Na2HPO4-KH2PO4(PBS) buffer solution(pH=6.82), the electrochemical behavior of mitoxantrone was studied by linear-sweep voltammetry and cyclic voltammetry at a Pt/C ion implantation modified microelectrode. A sensitive reduction peak was observed. The peak potential was -0.72 V(vs.SCE), the peak current was proportional to the concentration of mitoxantrone within the ranges of 7.0×10-8-9.0×10-7 mol/L and 1.0×10-6-2.4×10-5 mol/L, with a detection limit of 4.0×10-8 mol/L. The linear correlation coefficients were 0.9994 and 0.9992, respectively. This method has been applied to the direct determination of mitoxantrone in simulated urine. The recoveries were in the range from 96.2% to 105.9%. The reduction process was a quasi-reversible one with adsorptive characteristics at the Pt/C microelectrode. The electrode reaction rate constant ks and the electron transfer coefficient α of the system were determined to be 4.5 and 0.65 s-1, respectively. The experiments showed that Pt element had surely been implanted into the surface of the carbon fiber, and the atomic Pt improved the electrocatalytic activity. The Pt/C microelectrode had a good stability and reproducibility.     

STUDIES ON THE POLAROGRAPHIC ADSORP-TIVE COMPLEX WAVE OF RARE EARTH-ERIOCHROME AZUROL B (Ⅱ)

The rare earth ions form complexes with ECAB in 0.1 mol/L NaAc-HAc supporting electrolyte at pH 6. The composition of the complex was determined by spectrophotometric method to be 1: 2. The decrease of SAC peak height of ECAB is proportional to the concentration of rare earth ions in the range of 1×10~(-6)-2×10~(-5) mol/L. The reduction mechanism of ECAB and RE-ECAB is proposed based on the experimental evidence. The complexing group of ECAB with RE ions iust is the redox group, so when RE ions are added, there is no new peak appearing in the polarograms:, the only phenomenon observed is that the peak current of ECAB decreases. The fast protonation of EGAB carbanion formed after the second electron transfer results in the formation of colourless tetrahedral molecules.     

Potentiometric determination of diclofenac in pharmaceutical formulation by membrane electrode based on ion associate with base dye

The characteristics,performance and application of membrane electrode based on ion associate of diclofenac with base dye Safranine T are described.The electrode response to diclofenac has the sensitivity of 47±1.0 mV decade~(-1)over the range of 5×10~(-5)to 5×10~(-2)mol/L at pH 6-12,and the detection limit of 3.2×10~(-5)mol/L.The electrode is easy assembled at a relatively low cost has fast response time(2-4 s)and can be used for a period up to 3.5 months without any considerable divergence in potential.The proposed sensor displayed good selectivity for diclofenac in the presence of different substances.It was used to determine diclofenac in pharmaceuticals by means of the standard addition method.     

Flow-injection Determination of Cysteine,N-Acetyl Cysteine and Glutathione in Pharmaceuticals via Potassium Ferricyanide-Fe(Ⅲ) Spectrophotometric System

A simple flow injection spectrophotometric method is reported for the determination of cysteine,N-acetyl cysteine and glutathione based on the reduction of Fe(Ⅲ)/ferricyanide,the in situ reduced ions are reacted with unreduced portion of ferricyanide/Fe(Ⅲ) to form soluble Prussian blue,which is monitored at 735 nm.The calibration graphs are linear in the concentration ranges of(1―100)×10-6 mol/L for cysteine and N-acetyl cysteine,and(1―50)×10-6 mol/L for glutathione.The relative standard deviations of 1.8%,2.5% and 1.9% were found for eleven replicate analyses of 5×10-6 mol/L cysteine,N-acetyl cysteine and glutathione.The limits of detection(3σ blank) at 5×10-7 mol/L for cysteine,and 3×10-7 mol/L for N-acetyl cysteine and glutathione were obtained.The proposed method allowed 60 injections/h.The effects of common substances present in pharmaceuticals and human physiological fluids were examined.The method was applied to determining cysteine in pharmaceutical formulations with the recoveries in a range of 97% to 106% and the results obtained are agreed well with labeled values.     

Dual-modal Colorimetric and Fluorometric Method for Glucose Detection Using MnO2 Sheets and Carbon Quantum Dots

A novel dual-modal fluorometric and colorimetric method was developed for glucose detection using MnO2 sheets and carbon quantum dots(CQDs). The glucose could be oxidized by glucose oxidase, in accompanied witli the fbnnation of H2O2 intennediate, which resulted in the decomposition of MnO2 sheets, as well as tlie MnO2 sheets(brown) changed to Mn^2+ ions(colorless), which induced the absorption of MnO2 sheet decreased and the fluorescence of CQDs increased, consequently. The linear detection ranges of glucose are 5-1000 μmol/L by fluorescent method and 5-60 μmol/L by colorimetric method. The limits of detection of these two measurements are 2.11 and 2.18 μmol/L, respectively. This method is easy to conduct, has reasonable sensitive and selectivity, and could be applied for the glucose detection in real human senim.     

A POLAROGRAPHIC CATHODIC WAVE OF VITAMIN C

The cathodic wave of vitamin C has not been reported in the literature so far.Its E_(1/2)wasfound at-1.85 volt vs.Ag/AgCl(25℃)in the base solution of 0.1mol/L Bu_4NI.Variouselectrochemical and spectral techniques were used for studying the electrode reaction.They all showed that the electrode reaction was quasi-reversible while C was more than1.0×10~(-6)mol/L and mainly adsorptive while C was less than 1.0×10~(-6)mol/L at a mercuryelectrode.The electron transfer number was 2 and the electron transfer coefficient 0.57.Using DPP techniques,the linear response range was 5.0×10~(-7)-2.0×10~(-3)mol/L and thedetection limit 1×10~(-7)mol/L.     

Studies on the Electrochemical Behavior of Bleomycin at a Co/GC Ion Implantation Modified Electrode and Its Application

In a 0. 10 mol/L HAc-NaAc buffer solution (pH = 4. 59), a sensitive reduction peak of bleomycin was observed by linear sweep voltammetry at a Co/GC ion implantation modified electrode. The peak potential was-0. 73 V(iw. SCE). The peak current was proportional to the concentration of bleomycin over the range of 5.0 × 10-8-1.0× 10-6 and 1.0× 10-6-1. 0 × 10-5 mol/L with a detection limit of 2.0 × 10-8 mol/L. The electrochemical behavior of the reduction peak of bleomycin at the Co/GC modified electrode was studied by linear sweep and cyclic voltammetry and applied to the determination of bleomycin in urine. This method is simple, rapid and reliable. The reduction process is quasi-reversible. The experiments of AES and XPS showed that Co was surely implanted into the surface of GCE and the depth distribution of Co was in good agreement with Gooses normal distribution; the implanted Co at GCE improved the electrocatalytic activity.     

CHARACTERIZATION OF MOLECULAR MASS OF SIX WATER-SOLUBLE POLYSACCHARIDE -PROTEIN COMPLEXES FROM GANODERMA TSUGAE MYCELIUM

Six water-soluble polysaccharide-protein complexes coded as GM1, GM2, GM3, GM4, GM5 and GM6 wereisolated from the mycelium of Ganoderma tsugae by extracting with 0.2 mol/L phosphate buffer solution at 25, 40 and80℃, water at 120℃, 0.5 mol/L aqueous NaOH solution at 25 and 65℃, consecutively. Their chemical components wereanalyzed by using IR, GC, HPLC and ~(13)C-NMR, and some new results were obtained. The four samples GM1, GM2, GM3and GM4 are heteropolysaccharide-prote in complexes, in which, α- (1→3) linked D-glucose is the major monosaccharidewhile galactose, mannose and ribose are the secondary ones. GM5 and GM6 are β-(1→3)-D-glucan-protein complexes. Theprotein content increased from 32% to 69% with the progress of isolation. Weight-average molecu1ar mass M_w and theintrinsic viscosity [η] of the GM samples in 0.5 mol/L aqueous NaCl solution at 25℃ were measured systematically by laserlight scartering (LLS), size exclusion chromatography (SEC) combined with LLS, and viscometry. The M_w of GM1 to GM6are 35.5, 46.8, 58.9, 41.6, 3.3 and 22.0×10~4, respectively. The conformation and molecular mass of the two fractions of sample GM5 were characterized satisfactorily by SEC-LLS without further fractionation.