Immature mouse uterine tissue in organ culture: Estrogen-induced growth,morphology and biochemical parameters

Summary Although estrogens have been shown to stimulate a variety of morphologic and biochemical changes in the uterus in vivo, no clear consistent demonstration of similar responses in vitro have been made; thus, a defined organ culture system using the immature mouse uterus was established to study the possibility of demonstrating estrogenic responses in vitro. Uterine tissue from immature outbred mice (17 to 24 days of age) were cut crosswise in 1-mm³ coins and cultured in a defined medium in the absence of serum, phenol red, or growth factor supplements. Diethylstilbestrol (DES), a synthetic estrogen, was added to the media at doses ranging from 1 to 100 ng/ml. The effect of DES on uterine cell proliferation was assessed by morphologic changes in uterine epithelial and stromal cells, increase in number of epithelial cells per unit basement membrane, increase in height of luminal epithelial cells, and [³H]thymidine incorporation. Functional changes were determined by measuring the amounts of the estrogen-inducible uterine protein, lactoferrin, that was localized in the epithelial cells and secreted into the media, and the localization of the estrogen receptor in the cultured tissues. Results indicate that under the described conditions of culture, estrogens like DES can induce morphologic and biochemical responses in the uterus that are similar to those seen in vivo. This organ culture system will aid in the investigation of various mechanisms involved in the hormonal regulation of growth and differentiation of estrogen target tissues.     

An electrogenic proton pump in plasma membranes from the cellular slime mould Dictyostelium discoideum

Plants and fungi possess an outwardly directed plasma membrane proton pump that may regulate intracellular pH. We provide the first demonstration that amoebae of the slime mould Dictyostelium discoideum also possess a similar proton pump. It can be assayed either as an ATPase activity in highly purified plasma membranes or as a proton pump, after solubilization and reconstruction into liposomes. The pump is inhibited by vanadate, diethylstilbestrol (DES) and miconazole but not by azide or ouabain. The proton pump described here may represent the target for the action of DES and miconazole, both of which have previously been shown to induce stalk cell formation during the in vitro development of Dictyostelium.     

Mutagenic effectiveness and efficiency of EMS,DES and gamma-rays in rice

Summary Data on chlorophyll mutation frequency after treatment with EMS, DES and gamma-rays and sequential administration of gamma-rays and the two alkylating agents in three varieties of rice have been used to work out quantitatively the effectiveness and efficiency of each mutagen and combination treatment. For effectiveness, the order is EMS > DES and for efficiency it is EMS > DES > gamma-rays. In some sequential treatments (Gamma-rays + DES in IR8 and Basmati; DES + gamma-rays in IR8 and Jhona; Gamma-rays + EMS in IR8 and Basmati; and EMS + gamma-rays in IR8, Jhona and Basmati) mutation frequency is more than additive (synergistic) but these treatments are decisively less efficient because of their relatively high injurious effects in the M₁. generation. EMS induces more albinas than gamma-rays do. The mutational spectrum patterns induced by gamma-rays and DES are alike. In general, combination treatments tend to increase the frequency of albinas over other types of chlorophyll mutants.     

The mutagenicity and DNA-damaging activity of cyclic aliphatic sulfuric acid esters.

The cyclic aliphatic sulfuric acid esters 1,2-ethylene sulfate (ESF), 1,3-propylene sulfate (PSF) and 1,3-butylene sulfate (BSF) have been tested for their mutagenic and DNA-damaging activity. Mutagenicity of the compounds was established with his-auxotrophic indicator strains of Salmonella typhimurium using the in vitro plate test and the host-mediated assay technique with mice as host animals. The DNA-damaging activity was tested in a repair test with Proteus mirabilis mutants defective in DNA repair.In the repair test with a set of P. mirabilis strains (PG713 hcr^?rec^?: PG273 hcr⁺rec⁺) PSF and BSF showed a preferential growth inhibition of the repair-defective strain suggesting DNA-damaging activity of these chemicals. No such activity was found for ESF using the same concentrations of 5 and 15 μmol/plate.All cyclic sulfates revert the tester strain TA1535 of S. typhimurium in vitro indicating their ability to induce base substitutions. Compared with the reference compounds dimethyl sulfate (DMS), diethyl sulfate (DES), 1,3-propane sulfone (PPS) and 1,4-butane sulfone (BTS) the mutagenic activity in the plate test can be described as follows: PPS > PSF > BSF > BTS > ESF > DES > DMS.Dose-response studies in the host-mediated assay with tester strain TA1950 of S. typhimurium as genetic indicator system revealed a linear dosedependency of mutagenic activity. For PPS and PSF the lowest effective dose (LED) has been established as 10 μmol/kg. The LED for BSF and BTS was 50 μmol/kg, DMS and DES were mutagenic in doses of 2500 μmol/kg, while ESF was only weakly mutagenic with a LED of 5000 μmol/kg.The dose-response studies in the host-mediated assay and the results obtained in the in vitro spot test demonstrate similarities in the mutagenic action of the cyclic sulfates PSF and BSF and the respective sulfones, while the stronger alkylating compound ESF was a weak mutagen both in vitro and in vivo.     

Induction of sister-chromatid exchanges by indirect mutagens/carcinogens in cultured rat hepatoma and esophageal tumor cells and in Chinese hamster Don cells co-cultivated with rat cells

2 rat cell lines originated from ascites hepatoma AH66-B and esophageal tumor R1 were examined for their inducibility of sister-chromatid exchanges (SCEs) after treatment with 14 kinds of indirect mutagens/carcinogens, including 6 amine derivatives, 4 azo compounds, 3 aromatic hydrocarbons and 1 steroid. Of the 14 chemicals tested, 2-acetylaminofluorene (AAF), butylbutanolnitrosamine (BBN), dimethylnitrosamine (DMN), cyclophosphamide (CP), urethane, 2-methyl-4-dimethylaminoazobenzene (2-MeDAB), 3′-methyl-4-dimethylaminoazobenzene (3′-MeDAB), 4-o-tolylazo-o-toluidine (4-TT), benzo[a]pyrene (BP), 7,12-dimethyl-benz[a]anthracene (DMBA) and diethylstilbestrol (DES) were estimated to be effective inducers of SCEs in AH66-B and/or R1 cells, without the use of exogenous activating systems. Cell-mediated SCE tests with 6 selected chemicals, CP, 2-MeDAB, 4-TT, BP, DMBA and DES, showed a significant increase of SCEs in Chinese hamster Don-6 cells co-cultivated with AH66-B or R1 cells, depending on the number and sensitivity of AH66-B or R1 cells, as well as on the dose of chemicals tested, whereas singly cultured Don-6 cells were much less sensitive or almost insensitive to these chemicals. The above findings suggest that AH66-B and R1 cells may retain metabolic activities to convert a wide range of indirect mutagens/carcinogens into their active forms to induce SCEs, and that these cell lines provide simple and reliable screening systems in vitro, including the cell-mediated SCE assay, for detection of genotoxic agents, without the use of exogenous activation systems.     

Mutagenesis at the ad-3A and ad-3B loci in haploid UV-sensitive strains of Neurospora crassa. IV. Comparison of dose-response curves for MNNG, 4NQO and ICR-170 induced inactivation and mutation-induction

The genetic effects of MNNG, 4NQO and ICR-170 have been compared on 5 different UV-sensitive strains and a standard wild-type strain of Neurospora crassa with regard to inactivation and the induction of forward-mutations at the ad-3A and ad-3B loci. Whereas all UV-sensitive strains (upr-1, uvs-2, uvs-3, uvs-5 and uvs-6) are more sensitive to inactivation by MNNG and ICR-170 than wild-type, only uvs-5 shows survival comparable to wild-type after 4NQO treatment, all other strains are more sensitive to 4NQO. In contrast to the effects on inactivation, a wide variety of effects were found for the induction of ad-3A and ad-3B mutations: higher forward-mutation frequencies than were found in wild-type were obtained after treatment with MNNG or 4NQO for upr-1 and uvs-2, no significant increase over the spontaneous mutation frequency was found with uvs-3 after MNNG, 4NQO or ICR-170 treatment; mutation frequencies comparable to that found in wild-type were obtained with uvs-6 after MNNG, 4NQO or ICR-170 treatment and with upr-1 after ICR-170 treatment. Lower forward-mutation frequencies than were found in wild-type were obtained with uvs-2 after ICR-170 treatment and with uvs-5 after MNNG, 4NQO or ICR-170 treatment. These data clearly show that the process of forward-mutation at the ad-3A and ad-3B loci is under genetic control by mutations at other loci (e.g. upr-1, uvs-2, uvs-3, uvs-5 and uvs-6) and that the effect is markedly mutagen-dependent.     

Abscisic acid-triggered guard cell l-cysteine desulfhydrase function and in situ hydrogen sulfide production contributes to heme oxygenase-modulated stomatal closure

Recent studies have demonstrated that hydrogen sulfide (H₂S) produced through the activity of l -cysteine desulfhydrase (DES1) is an important gaseous signaling molecule in plants that could participate in abscisic acid (ABA)-induced stomatal closure. However, the coupling of the DES1/H₂S signaling pathways to guard cell movement has not been thoroughly elucidated. The results presented here provide genetic evidence for a physiologically relevant signaling pathway that governs guard cell in situ DES1/H₂S function in stomatal closure. We discovered that ABA-activated DES1 produces H₂S in guard cells. The impaired guard cell ABA phenotype of the des1 mutant can be fully complemented when DES1/H₂S function has been specifically rescued in guard cells and epidermal cells, but not mesophyll cells. This research further characterized DES1/H₂S function in the regulation of LONG HYPOCOTYL1 (HY1, a member of the heme oxygenase family) signaling. ABA-induced DES1 expression and H₂S production are hyper-activated in the hy1 mutant, both of which can be fully abolished by the addition of H₂S scavenger. Impaired guard cell ABA phenotype of des1/hy1 can be restored by H₂S donors. Taken together, this research indicated that guard cell in situ DES1 function is involved in ABA-induced stomatal closure, which also acts as a pivotal hub in regulating HY1 signaling.     

UV与DES复合诱变选育高产脂肪酶菌株

以产脂肪酶菌株BaciUus sp CS-4为出发菌株,进行了UV与硫酸二乙酯(DES)复合诱变处理.筛选出一株高酶活的目的菌株,命名为Bacillus spDE-8.其酶活为每毫升14.85U,比出发菌株提高48.2%.传代实验证明,其遗传性能稳定.     

已烯雌酚对免疫性卵巢早衰小鼠卵巢细胞凋亡的影响

目的探讨已烯雌酚对免疫性卵巢早衰小鼠卵巢颗粒细胞和卵母细胞凋亡的影响。方法以小鼠透明带3蛋白的第330～342个氨基酸的序列（NSSSSQFQIHGPR）合成透明带多肽并作免疫原,免疫SPF级BALB/c雌性小鼠。设对照、模型和已烯雌酚组。灌胃给药4周后,采用Tunel方法检测颗粒细胞与卵母细胞的细胞凋亡数并作统计学分析。结果已烯雌酚组可显著减少小鼠卵巢卵母细胞的凋亡率（P〈0.05）。结论已烯雌酚组能改善免疫性卵巢早衰小鼠体重降低症状并能诱导发情。已烯雌酚能改善免疫性卵巢早衰小鼠症状的机制可能是由于能有效地抑制卵母细胞的凋亡所引起的。