Inhibition of gap junctional intercellular communication in heptachlor- and heptachlor epoxide-treated normal human breast epithelial cells

Based on the concern of organochlorides in the environment and in human tissue, this study was designed to determine whether various noncytotoxic levels of heptachlor and heptachlor epoxide could inhibit, reversibly, gap junctional intercellular communication in human breast epithelial cells (HBEC). Cytotoxicity and gap junctional intercellular communication (GJIC) were evaluated by lactate dehydrogenase assay and fluorescence redistribution after photobleaching analysis, respectively. Both heptachlor and heptachlor epoxide were noncytotoxic up to 10 μg/ml. At this concentration, heptachlor and heptachlor epoxide inhibited GJIC of normal human breast epithelial cells after 1 h treatment. Within a 24 h treatment with heptachlor and heptachlor epoxide at 10 μg/ml, recovery of GJIC had not returned. GJIC completely recovered after a 12 h treatment of 1 μg/ml heptachlor epoxide, but it did not recover after a 24 h treatment of 1 μg/ml heptachlor. RT-PCR and Western blots were analyzed to determine whether the heptachlor or heptachlor epoxide might have altered the steady-state levels of gap junction mRNA and/or connexin protein levels or phosphorylation state. No significant difference in the level of connexin 43 (Cx43) message between control and heptachlor-treated cells was observed. Western blot analyses showed hypophosphorylation patterns in cells treated with 10 μg/ml heptachlor and heptachlor epoxide for 1 h with no recovery within 24 h. Immunostaining of Cx43 protein in normal HBEC indicated that heptachlor and heptachlor epoxide caused a loss of Cx43 from the cell membranes at noncytotoxic dose levels. Taken together, these results suggest that heptachlor and heptachlor epoxide can alter GJIC at the post-translational level, and that, under the conditions of exceeding a threshold concentration in the breast tissue containing ‘initiated’ cells for a long time and not being counteracted by anti-tumor-promoting chemicals, they could act as breast tumor promoters.     

Stochastic versus deterministic variability in simple neuronal circuits: II. Hippocampal slice.

Long time series of Schaffer collateral to CA1 pyramidal cell presynaptic volleys (stratum radiatum) and population spikes (stratum pyramidale) were evoked (driven) in rat hippocampal slices. From the driven CA1 region in normal [K+] perfusate, both population spike amplitude and an input-output function consisting of population spike amplitude divided by the presynaptic volley amplitude were analyzed. Raising [K+] in the perfusion medium to 8.5 mM, slices were induced to spontaneously burst fire in CA3 and long time series of inter-burst intervals were recorded. Three tests for determinism were applied to these series: a discrete adaptation of a local flow approach, a local dispersion approach, and nonlinear prediction. Surrogate data were generated to serve as mathematical and statistical controls. All of the population spike (6/6) and input-output (6/6) time series from the normal [K+] driven circuitry were stochastic by all three methods. Although most of the time series (5/6) from the autonomously bursting high [K+] state failed to demonstrate evidence of determinism, one (1/6) of these time series did demonstrate significant determinism. This single instance of predictability could not be accounted for by the linear correlation in these data.     

Intermediates of adeno-associated virus DNA replication in vitro.

Intermediates of adeno-associated virus type 2 (AAV) DNA replication in an in vitro assay have been characterized. The assay involves rescue and replication of an AAV insert in pBR322. Intermediates were shown to be duplex molecules in which at least one terminus was in the extended configuration, in contrast to the hairpinned ends seen after rescue in the absence of AAV DNA replication. Also present were linear double-stranded dimers, which were characterized as either head-to-head or tail-to-tail tandems; no head-to-tail dimers were detected. The results are in accord with the current model of AAV DNA replication.     

Increased shikonin production by hairy roots of Lithospermum erythrorhizon in two phase bubble column reactor

Summary Plant hairy root cultures of Lithospermum erythrorhizon were carried out to produce shikonin derivatives by employing in situ extraction with n-hexadecane in a shake flask and a bubble column bioreactor. Over 95 % shikonin produced was recovered in the n-hexadecane layer. In flask cultures the maximum concentration of shikonin with n-hexadecane extraction was 3 times higher than that obtained without extraction. In the two phase bubble column reactor, 572.6 mg/L of shikonin and 15.6 g/L of dry cell mass were obtained after 54 days. Shikonin was produced at a constant level of 10.6 mg/L day during this period.     

零下低温对杂交杨树皮层膜脂组成的影响

以不耐寒的美洲黑杨（Ｐｏｐｕｌｕｓｄｅｌｔｏｉｄｅｓｃｖ．“Ｌｕｘ”Ｉ－６９／５５,父本）和耐寒性较强的欧美杨（Ｐ．ｅｕｒａｍｅｒｉｃａｎａｃｌｃｖ．Ｉ－４５／５１,母本）的４个杂交Ｆ＿１代无性系（９５杨、５５９杨、６００杨和１３８１杨）为材料,分析了零下低温寒潮前后枝条皮层的脂质组成。结果表明,寒潮影响下,皮层中磷脂含量增加而组成基本不变,膜脂脂肪酸组成的变化规律是：寒潮前脂肪酸不饱和指数（ＩＵＦＡ）值大的无性系,寒潮前后的ＩＵＦＡ值变化量小;寒潮前ＩＵＦＡ值较小的无性系,寒潮前后ＩＵＦＡ值变化量较大。本文借用力学概念,提出相对抗性概念,给出杨树无性系的相对抗性序列。序列表明Ｆ＿１代无性系的耐寒性已较不耐寒的父本提高,这与田间观察基本一致。     

Absence of a direct role for RNase HI in initiation of DNA replication at the oriC site on the Escherichia coli chromosome.

On the basis of the experiments carried out with rnhA224 mutants, we previously concluded that RNase HI is not essential for initiation of Escherichia coli chromosome replication at oriC (T. Kogoma, N.L. Subia, and K. von Meyenburg, Mol. Gen. Genet. 200:103-109, 1985). In light of the recent finding that rnhA224 is a UGA nonsense mutation which can be leaky in certain genetic backgrounds, we reexamined this conclusion with the use of rnhA339 (Null)::cat mutants. The possibility that recB+ is required for initiation at the alternative origins (oriKs) of replication in rnhA mutants was also tested. The results clearly indicated that RNase HI is not essential for oriC initiation and that recB+ is not required for initiation at oriK sites.