Hyperinsulinemic yellow KK mice and norepinephrine turnover

To clarify whether hyperinsulinemia accelerates sympathetic nervous system (SNS) activity, norepinephrine (NE) turnover, a reliable indicator of SNS activity, was measured in the interscapular brown adipose tissue (IBAT) and heart of hyperinsulinemic yellow KK and normoinsulinemic C57BL control mice at 12 weeks of age. The yellow KK mice were more obese and had higher levels of plasma glucose (about 2.3 times) and of plasma insulin (about 24 times) than did the control mice. In IBAT, the rate of NE turnover following blockade of NE synthesis with alpha-methyl-p-tyrosine (alpha-MPT) was significantly slower in yellow KK mice than in C57BL mice, although in heart, no significant difference between both groups was observed in NE turnover. These results suggest that hyperinsulinemia dose not always increase NE turnover, and furthermore that the reduced NE turnover in IBAT of yellow KK mice may be one of the important factors in the development of obesity of this animal, as it is recognized that brown adipose tissue is a main effector of diet-induced thermogenesis and its defect or absence would predispose to obesity.     

Biologically active phenolics from Lilium longiflorum

The new phenolics, the trans-p-coumaroyl and the feruloyl esters of 2,3-dihydroxy-1,2-propanedicarboxylic acid and itaconic acid, were isolated from Lilium longiflorum; the cis-p-coumaroyl ester was also detected. The biological activities of the trans-p-coumaroyl ester are described.     

A DNA polymorphism from five inbred strains of the mouse identifies a functional class of domesticus-type Y chromosome that produces the same phenotypic distribution of gonadal hermaphrodites

The wild-derived CLA inbred strain of the house mouse contains a domesticus-type Y chromosome that lacks a 2.3-kb TaqI band with fragment 1 of the AC11 probe. The CLA Y chromosome also causes a low frequency of XY gonadal hermaphrodites when backcrossed to the C57BL/6J strain (F. G. Biddle and Y. Nishioka, 1988. Genome, 30:870-878). A similar domesticus-type Y chromosome, lacking the 2.3-kb TaqI band has now been found in the four historical inbred strains AKR/J, MA/MyJ, PL/J, and RF/J. When backcrossed to C57BL/6J, these four Y chromosomes cause low frequencies of gonadal hermaphrodites similar to the CLA Y and phenotypic distribution of types of gonad are indistinguishable from that with the CLA Y. The absence of the 2.3-kb TaqI band appears to be a polymorphism among domesticus-type Y chromosomes that identifies one of the three functional classes that, so far, can be distinguished only by their effects on testis differentiation in backcross test fetuses with the C57BL/6J strain. Three other historical inbred strains, BUB/BnJ, ST/bJ, and SWR/J, with a domesticus-type Y chromosome but containing the 2.3-kb TaqI band, were also assayed. They permit normal testis development in backcross test fetuses with C57BL/6J.     

Protein S is essential for the activated protein C-catalyzed inactivation of platelet-associated factor Va

The prothrombin-converting activity of Factor Xa was enhanced by thrombin-stimulated Factor V-deficient platelets and supplementary extraneous Factor Va, and also by thrombin-stimulated normal human platelets. Both extraneous Factor Va and intra-platelet Factor Va were equally inactivated by a gamma-carboxyglutamic acid-containing plasma protease, activated protein C. However, a relatively larger amount of activated protein C was required for efficient inactivation of platelet-associated Factor Va as compared with the amount of activated protein C needed for inactivation of phospholipid vesicle-associated Factor Va. Protein S, another gamma-carboxyglutamic acid-containing plasma protein, increased the rate of the inactivation of platelet-associated Factor Va about 25-fold. This stimulating effect was observed only slightly with the thrombin-modified protein S. Thus, it was concluded that protein S is essential for the process of inactivation of platelet-associated Factor Va by activated protein C.     

Structure and expression of human thrombomodulin, a thrombin receptor on endothelium acting as a cofactor for protein C activation.

We have deduced the entire 575-amino acid sequence of the human thrombomodulin precursor from cDNA clones. The precursor starts with an 18-residue signal peptide domain, followed by the NH2-terminal domain, a domain with six epidermal growth factor-like structures, an O-glycosylation site-rich domain, a 24-residue transmembrane domain and a cytoplasmic domain. Simian COS cells transfected with the expression vector pSV2 containing thrombomodulin cDNA synthesized immunoreactive and functionally active thrombomodulin.     

Evolution of a mouse Y chromosomal sequence flanked by highly repetitive elements

Mammalian primary sex is determined by the presence or absence of the Y chromosome. However, little is known about the molecular processes through which the Y chromosome exerts its action. We applied recombinant DNA techniques to isolate mouse Y chromosomal fragments and described previously a clone designated as AC11 (Y. Nishioka and E. Lamothe. 1986. Genetics, 113:417-432). To obtain information on DNA sequences that flank AC11, we screened a mouse genomic library for the presence of AC11-related sequences and isolated over 50 positive clones. In this report we describe clones ACC2 and ACC3, both of which contain highly repetitive elements. Using a male-specific portion of these clones, we compared DNA's isolated from mice (Mus musculus, M. hortulanus, M. spretus, M. cookii, M. pahari, and M. platythrix), rat, hamster, and guinea pig and obtained results that agree with the phylogenetic relationships deduced from morphological and biochemical studies. The male-specific accumulation of the related sequences was found only in M. musculus, M. hortulanus, and M. spretus.     

Reduced brown adipose tissue thermogenesis of obese rats after ovariectomy

Brown adipose tissue (BAT) thermogenesis was assessed by measuring mitochondrial guanosine diphosphate (GDP) binding, cytochrome oxidase activity and oxygen consumption in ovariectomized (OVX) and sham-operated rats. The food intake and body weight of OVX rats increased more than those of controls and OVX rats became obese. Mitochondrial GDP binding, as an indicator of thermogenic activity, cytochrome oxidase activity, as a marker of mitochondrial abundance, and mitochondrial respiration of BAT in OVX rats were significantly reduced compared with those in controls. And, also, even when OVX rats were restricted in food intake (pair-gained) to produce comparable changes in body weight with sham-controls, or matched in food intake (pair-fed) with sham-controls, these parameters in both pair-gained and pair-fed OVX groups were decreased markedly compared to those in sham-controls. As expected, body weight in pair-fed OVX rats increased significantly more than that in sham-controls. In response to cold exposure, these parameters of OVX rats increased as much as those of controls did. These results suggest that reduced brown adipose tissue thermogenesis might be one of the important factors that are responsible for the development of obesity after OVX.     

Localization of bindin expression during sea urchin spermatogenesis

Expression of the bindin gene was examined in testicular cells of the sea urchin Strongylocentrotus purpuratus. In situ hybridization studies, using an 35S-labeled antisense RNA probe transcribed from a bindin cDNA, reveal that bindin mRNAs are localized in spermatogenic cells displaced towards the lumens of maturing testicular acini. Little or no hybridization is observed in spermatogenic cells displaced towards the perivisceral epithelium or in somatic cells of the testis. A similar localization of the bindin protein itself is observed using a rhodamine-conjugated polyclonal antibody against bindin, which shows a punctate immunofluorescence pattern in late spermatogenic cells. Immunogold labeling of ultrathin sections and electron microscopy reveal that this punctate immunofluorescence is an apparent result of localized deposits of bindin in intracellular vesicles. Through the terminal stages of spermatogenesis, these bindin-containing vesicles apparently fuse to form the single acrosomal vesicle of the mature spermatozoon. These results indicate 1) that bindin mRNAs are transcribed relatively late in spermatogenesis, 2) that bindin is translated soon after production of its mRNA, 3) that bindin quickly associates with intracellular vesicles during or soon after its synthesis, and 4) that these vesicles fuse to form the single acrosomal vesicle during the terminal stage of spermatogenesis.     

Changing localizations of site-specific sperm surface antigens during sea urchin fertilization

Indirect immunofluorescence studies show that monoclonal antibody (mAb) J18/2 binds site-specifically to surface antigens localized over the acrosome and tail regions of mature Strongylocentrotus purpuratus spermatozoa. Within 5 min after induction of the acrosome reaction by exposure to egg jelly or ionophore A23187, these surface antigens become detectable over the lateral region of the head so that the entire surface of the spermatozoon is labeled. Polyspermically fertilized S. purpuratus eggs fixed at varying times after insemination and exposed to mAb J18/2 reveal that these surface antigens are quickly incorporated into the egg plasma membrane and begin to disperse as early as 1.5 min after insemination. At subsequent times, they undergo further dispersal so that by 45 min they are distributed over the entire surface of the egg. These results suggest that the sperm surface components recognized by mAb J18/2 gain the ability to disperse laterally during the acrosome reaction and proceed to do so in the egg plasma membrane after fertilization.