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An underlying assumption of laboratory-based toxicity tests is that the sensitivity of organisms in the laboratory (in vitro) is comparable to that in the field (in situ). We tested this assumption by exposing estuarine amphipods (Chaetocorophium cf. lucasi) to a concentration series of cadmium-spiked sediments in vitro and in situ for 10 d. In situ exposures were conducted within plastic-mesh cages on an intertidal mudflat. To characterize exposure, we. measured interstitial water cadmium concentrations OWCO, acid volatile sulfide (AVS), and simultaneously extracted Cd (SEMcd) at the beginning and end of the exposures. Between day 0 and day 10, AVS decreased in both in vitro and in situ exposures, while IWcd levels declined less in vitro (median 27%) than in situ (median 76%). Despite more extreme conditions of temperature (10-36 deg.C) and salinity (18-22%o) in situ. in vitro and in situ exposures showed comparable survival responses based on SEMcd/AVS (LC50 [95% CI]: 1.6 [1.46-1.78] and 1.8 [1.761.83], respectively), with the onset of marked mortality above a SEMcd/AVS value of aboutone and minimal survival (<5%) above a value of two. Based on IWcd concentrations, however, sensitivity was significantly greater in vitro (LC50 = 0.41 mg Cd/L [0. 1710.959], in situ LC50 = 1.6 mg Cd/L [1.15-2.16]). We concluded that, in our tests, amphipod sensitivity in vitro was equal to or greater than its sensitivity in situ. [ABSTRACT FROM AUTHOR]

Sediment physicochemical characteristics, benthic community structure, and toxicity were measured at reference and contaminated intertidal mudflats around the North Island of New Zealand. Chronic whole-sediment toxicity tests were conducted with the estuarine amphipod Chaetocorophium lucasi and the marine bivalve Macomona liliana, and pore-water toxicity tests were conducted with embryos of the echinoid Fellaster zelandiae. Although concentrations of organic chemicals and heavymetals were up to several orders of magnitude higher at the sites considered to be contaminated, levels of contamination were relatively low compared to internationally based sediment quality guidelines. Although no pronounced difference was found in benthic community structure between reference and contaminated sites, multivariate analysis indicated that natural sediment characteristics (total organic carbon,acid-volatile sulfide, ammonium) and factors related to contamination (dichlorodiphenyltrichloroethane, sediment toxicity) may have been affecting community structure. Although benthic effects caused by present levels of contamination are not yet dramatic, subtle changes in community structure related to pollution may be occurring. The two whole-sediment and the pore-water toxicity tests presented different response patterns. Growth of C. lucasi and M. liliana was a less sensitive endpoint than survival. None of the three toxicity tests responded more strongly to the contaminated than to the reference sites, thatis, neither natural-sediment and pore-water characteristics nor unmeasured contaminants affected the test organisms. It is possible that sediment collection and handling may have induced chemical changes, confounding interpretation of toxicity tests. [ABSTRACT FROM AUTHOR]

Objective To assess the rates, timing and causes of neonatal deaths and the burden of stillbirths in rural Uttar Pradesh, India. We discuss the implications of our findings for neonatal interventions. Methods We used verbal autopsy interviews to investigate 1048 neonatal deaths and stillbirths. Findings There were 430 stillbirths reported, comprising 41% of all deaths in the sample. Of the 618 live births, 32% deaths were on the day of birth, 50% occurred during the first 3 days of life and 71% were during the first week. The primary causes of death on the first day of life (i.e. day 0) were birth asphyxia or injury (31%) and preterm birth (26%). During days 1-6, the most frequent causes of death were preterm birth (30%) and sepsis or pneumonia (25%). Half of all deaths caused by sepsis or pneumonia occurred during the first week of life. The proportion of deaths attributed to sepsis or pneumonia increased to 45% and 36% during days 7-13 and 14-27, respectively. Conclusion Stillbirths and deaths on the day of birth represent a large proportion of perinatal and neonatal deaths, highlighting an urgent need to improve coverage with skilled birth attendants and to ensure access to emergency obstetric care. Health interventions to improve essential neonatal care and care-seeking behavior are also needed, particularly for preterm neonates in the early postnatal period. [ABSTRACT FROM AUTHOR]

Autocrine epidermal growth factor receptor activation by transforming growth factor alpha (TGF alpha) has been implicated in growth stimulation during epithelial neoplasia. Using keratinocytes isolated from mice with genetic defects in TGF alpha expression, we tested whether TGF alpha is required for transformation by the v-rasHa oncogene. Introduction of v-rasHa into primary epidermal cultures using a retroviral vector stimulated growth of both control (TGF alpha +/+, BALB/c) and TGF alpha-deficient (TGF alpha -/-, wa-1) keratinocytes. Moreover, v-rasHa elicited characteristic changes in marker expression (keratin 1 was suppressed; keratin 8 was induced), previously shown to be associated with epidermal growth factor (EGF) receptor activation, in both TGF alpha +/+ and TGF alpha -/- keratinocytes. v-rasHa markedly increased secreted (> 10-fold) and cell-associated (2-3-fold) TGF alpha levels in keratinocytes from TGF alpha +/+ and BALB/c mice, but not TGF alpha -/- or wa-1 mice. Based on Northern blot analysis, v-rasHa induced striking up-regulation of transcripts encoding the additional EGF family members amphiregulin, heparin-binding EGF-like growth factor, and betacellulin in cultured keratinocytes from all four mouse strains. Interestingly, in addition to the normal 4.5-kilobase TGF alpha transcript, wa-1 keratinocytes expressed two additional TGF alpha transcripts, 4.7 and 5.2 kilobases long. All three transcripts were up-regulated in response to v-rasHa, as well as exogenous TGF alpha or keratinocyte growth factor treatment, and were also detected in RNA isolated from wa-1 brain and skin. In vivo, v-rasHa keratinocytes from control as well as TGF alpha-deficient mice produced squamous tumors when grafted onto nude mice, and these lesions expressed high levels of amphiregulin, heparin-binding EGF-like growth factor, and betacellulin mRNA, regardless of their TGF alpha status. These findings indicate that TGF alpha is not essential for epidermal neoplasia induced by the v-rasHa oncogene and suggest that another EGF family member(s) may contribute to autocrine growth stimulation of ras-transformed keratinocytes.

The maturation of epidermal keratinocytes is a tightly regulated, stepwise process which requires protein kinase C (PKC) activation. We investigated the effect of elevated extracellular Ca2+, a potent differentiation signal which increases cellular sn-1,2-diacylglycerol levels, on the PKC isozyme profile of cultured murine keratinocytes. Five PKC isozymes (alpha, delta, epsilon, zeta, and eta) were detected by immunoblotting. During Ca(2+)-induced differentiation, total cellular PKC alpha decreased, PKC epsilon and eta increased 3-5-fold, and the level of other PKC isozymes was relatively unchanged. PKC alpha underwent a progressive translocation from the soluble to the particulate fraction following elevation of extracellular Ca2+. The kinetics of PKC alpha translocation corresponded with the induction of keratinocyte differentiation markers. Both PKC delta and epsilon were selectively lost from the soluble fraction of keratinocytes exposed to elevated extracellular Ca2+, resulting in an increase in the proportion of these isoforms in the particulate fraction. PKC eta increased in both the soluble and particulate fractions, while PKC zeta did not change in amount or distribution during keratinocyte differentiation. Selective down-regulation of PKC isoforms by either 12-deoxyphorbol-13-phenylacetate or bryostatin 1 inhibited Ca(2+)-induced expression of differentiation markers at doses most specific for the down-regulation of PKC alpha. Taken together, these observations suggest that the induction of keratinocyte differentiation by Ca2+ results in the activation of specific PKC isozymes.

Primary mouse keratinocytes expressing the v-rasHa oncogene (v-rasHa keratinocytes) produce squamous papillomas when grafted onto nude mice and respond abnormally to signals for terminal differentiation both in vivo and in vitro. Since protein kinase C (PKC) activators and v-rasHa induce similar phenotypic changes in cultured keratinocytes, and cellular diacylglycerol levels are constitutively elevated in ras-transformed keratinocytes, we tested whether PKC is a downstream target for oncogenic ras in this cell type. Ca(2+)-dependent PKC activity was increased in lysates from cultured v-rasHa keratinocytes when compared to control cells; in contrast, Ca(2+)-independent activity decreased. Similar to PKC activators, v-rasHa blocked Ca(2+)-mediated expression of the early epidermal differentiation markers keratins K1 and K10 while inducing aberrant expression of K8. Pretreatment of v-rasHa keratinocytes with bryostatin to block PKC function restored Ca(2+)-mediated expression of K1 and K10 and blocked abnormal expression of K8, suggesting that these responses are mediated by the PKC pathway. Furthermore, expression of K1 is restored at bryostatin doses which specifically down-modulate PKC-alpha, the only Ca(2+)-dependent PKC isozyme detected in cultured keratinocytes. In contrast to the inhibition of K1 and K10, Ca(2+)-induced expression of the late epidermal differentiation markers loricrin, filaggrin, and keratinocyte transglutaminase was accelerated by v-rasHa, as previously reported in normal keratinocytes treated with PKC activators. Pretreatment of v-rasHa keratinocytes with bryostatin blocked expression of late markers in these cells, and this response was correlated with down-regulation of PKC-alpha. The results of this study suggest that oncogenic ras alters keratinocyte differentiation by altering the function of the PKC signaling pathway, and that PKC-alpha is the specific isozyme involved in down-modulating expression of keratins K1 and K10 and up-regulating expression of loricrin, filaggrin, and keratinocyte transglutaminase.