Recovery of poly-3-hydroxybutyrate (PHB) in three chlorinated solvents with or without acetone pretreatment and degradation of extracted PHB (99% pure) in hot chloroform were studied. When lyophilized Alcaligenes eutrophus biomass was used, the best results were obtained with acetone pretreatment and solvent reflux for 15 min in methylene chloride or chloroform. Recovered PHB had a 95% purity and molecular weights (w) of 1,050,OOO and 930,000 g/mol respectively. Further heating resulted in a serious & loss at reflux temperatures. Degradation of extracted PHB at 110°C in chloroform was due to random and chain-end scission, the former being predominant.

A direct in vitro effect of 17p-estradiol Although gonadal steroids have a profound effect on skeletal tissues (1-3), and estrogen deficiency has been established as a major etiologic factor in postmenopausal osteoporosis (1, 2), their influence has been considered to be indirect. This prevailing opinion was due to a lack of evidence for either specific estradiol receptors in bone (4) or direct biologic effects of sex steroids on bone cells (5, 6). The situation has dramatically reversed since recent studies have demonstrated significant, albeit low, concentrations of 17/3-estradiol (E2) receptors (refs. 7-9; 1) as well as androgen receptors ( ¶) in bone cells. The brain type (BB) isoenzyme of creatine kinase (CK; ATP:creatine N-phosphotransferase, EC 2.7.3.2; ref. 10) involved in the "energy buffer" system, which regulates cellular concentrations of ATP and ADP, is the major component (11) of the E2-induced protein of the rat uterus (12). E2-induced protein synthesis, and more recently, modulation of CK activity, has been a useful marker for studies on the mechanism ofaction of E2 in uterus and in other tissues that contain E2 receptors (13), because of its rapid response to E2 in vivo and in vitro (14). Moreover, it is a convenient marker for estrogen-modulated gene expression, since E2 treatment increases the steady-state level of mRNA for CK BB in the rat uterus (15). The speed and sensitivity of the assay for CK activity makes CK stimulation an efficient response marker to detect the action of E2 and other hormones (16) in skeletal tissues. In this report, we present evidence that E2 acts directly on cultured osteoblasts and epiphyseal cartilage cells, leading to increased CK activity as well as increased [3H]thymidine incorporation into DNA. Moreover, we report a rapid and sex-specific action of E2 and testosterone (T) on these markers in bones of prepubertal rats. Rat epiphyseal cells were obtained from vitamin Ddeficient 16-to 18-day-old rats, which have a wider epiphyseal cartilage zone than normally fed rats. Epiphyseal cartilage plates were isolated under a binocular dissecting microscope. Cells released by digestion with 0.25% collagenase (Worthington) in phosphate-buffered saline for 60 min at 370C were cultured as described above for calvaria cells (in 2 mM Ca2+). tTo whom reprint requests should be addressed. ¶Spelsberg, T. MATERIALS AND METHODS

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Vascular placental insufficiency is considered a common pathogenic factor in human intrauterine growth retardation (IUGR), resulting in small-for-gestational-age, asymmetric newborns. IUGR neonates experience higher morbidity and mortality rates, as well as a possible contribution towards late sequelae, such as hypertension, and cardiovascular disease in adulthood. To simulate vascular placental insufficiency, an experimental rabbit IUGR model was used. Intrauterine growth retardation was achieved by ligation of 25-30% uteroplacental vessels of half of the fetuses during the last third of gestation. Ischemic fetuses were significantly small, asymmetric, and had a disproportionately small body with a relatively large head. The kidneys from all groups were analyzed for relative estimated glomeruli number (REGN) using an unbiased blind design. The glomeruli number was significantly reduced in the asymmetric IUGR rabbit fetuses, probably due to decreased renal vascular supply. Our results support the concept that the reduced number of glomeruli may contribute to impaired renal function, thus predisposing to neonatal renal dysfunction and late sequelae, such as adult hypertension. This study emphasizes the clinical importance of early IUGR diagnosis and prevention.

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