Sexually dimorphic gene expression in the developing mouse gonad

Over the course of a few days, the bipotential embryonic mouse gonad differentiates into either a testis or an ovary. Though a few gene expression differences that underlie gonadal sex differentiation have been identified, additional components of the testicular and ovarian developmental pathways must be identified to understand this process. Here we report the use of a PCR-based cDNA subtraction to investigate expression differences that arise during gonadal sex differentiation. Subtraction of embryonic day 12.5 (E12.5) XY gonadal cDNA with E12.5 XX gonadal cDNA yielded 19 genes that are expressed at significantly higher levels in XY gonads. These genes display a variety of expression patterns within the embryonic testis and encode a broad range of proteins. A reciprocal subtraction (of E12.5 XX gonadal cDNA with E12.5 XY gonadal cDNA) yielded two genes, follistatin and Adamts19, that are expressed at higher levels in XX gonads. Follistatin is a well-known antagonist of TGFbeta family members while Adamts19 encodes a new member of the ADAMTS family of secreted metalloproteases.     

Production of prostaglandins by fetal rat lung type II pneumonocytes and fibroblasts

The output of prostaglandins I2, E2, F2 alpha and 13,14-dihydro-15-keto-PGF2 alpha (PGFM) from third passage day 20 rat fetal fibroblasts and type II alveolar pneumonocytes was studied. In 2 h incubations, the output levels for each cell type were: PGI2 greater than PGE2 much greater than PGF2 alpha = PGFM when cells were incubated with Ca2+ ionophore A23187 (10 microM) or arachidonic acid (1 microgram/ml).     

Sexually dimorphic expression of myosin heavy chains in the adult mouse masseter.

Little is known regarding the role of androgenic hormones in the maintenance of myosin heavy chain (MHC) composition of rodent masticatory muscles. Because the masseter is the principal jaw closer in rodents, we felt it was important to characterize the influence of androgenic hormones on the MHC composition of the masseter. To determine the extent of sexual dimorphism in the phenotype of masseter muscle fibers of adult (10-mo-old) C57 mice, we stained tissue sections with antibodies specific to type IIa and IIb MHC isoforms. Females contain twice as many fibers containing the IIa MHC as males, and males contain twice as many fibers containing the IIb MHC as females. There is a modest amount of regionalization of MHC phenotypes in the mouse masseter. The rostral portions of the masseter are composed mostly of type IIa fibers, whereas the midsuperficial and caudal regions contain mostly type IIb fibers. Using immunoblots, we showed that castration results in an increase in the expression of type IIa MHC fibers in males. Ovariectomy has no effect on the fiber type composition in females. We conclude that testosterone plays a role in the maintenance of MHC expression in the adult male mouse masseter.     

Proliferation of type II pneumonocytes after X-irradiation

This paper reports preliminary data on the proliferative response of type II cells in the mouse lung over a five-month period after external thoracic doses of 2, 5, 10 and 12 Gy of X-rays. The DNA labelling index (LI) of control (0 Gy) mice was at all times exceedingly low (0.3-0.4 per cent). The LI after 2 and 5 Gy showed a slight though transient fall below controls during the first week post-irradiation, and thereafter the LIs were similar to the controls for the 5 months of the experiment. The LI after 10 and 12 Gy again showed a significant depression during the first week, but this was followed by a significant increase (P = 0.01) in LI which peaked at 4 weeks after irradiation. The LI returned to control values at 3-4 months and again rose significantly (P = 0.05) at 5 months. The first wave of proliferation corresponds to data showing an increase in surfactant in alveolar fluids within 2-6 weeks of 10-15 Gy of X-rays; and the second wave coincides with the pneumonitic phase and is consistent with a delay before the alveolar epithelial continuity is sufficiently compromised by the low rates of type I cell loss to trigger a compensatory wave of type II cell divisions. This relatively chronic radiation response is discussed and contrasted with the dramatic and immediate hyperplastic responses which many toxic irritants produce in type II epithelial cells.     

Sexually dimorphic expression of prepro-orexin mRNA in the rat hypothalamus

The neuropeptides orexin A and B are expressed in the lateral hypothalamic area and are involved in the regulation of energy homeostasis and arousal. Recent results showed gender differences in the expression of orexin receptor subtypes in rats. In the present study, we analyzed the mRNA expression of prepro-orexin (PPO) in the hypothalamus of male and female rats using quantitative real-time PCR. We found significantly higher levels of PPO mRNA in the hypothalamus of female rats compared to male rats. Our study indicates a sex-dependent regulation of hypothalamic PPO expression and suggests gender-specific functions of orexins.     

Lipopolysaccharide increases alveolar type II cell number in fetal mouse lungs through Toll-like receptor 4 and NF-kappaB

Chorioamnionitis is a major cause of preterm delivery. Infants exposed to inflammation in utero and then born preterm may have improved lung function in the immediate postnatal period. We developed a mouse model of chorioamnionitis to study the inflammatory signaling mechanisms that might influence fetal lung maturation. With this in vivo model, we found that Escherichia coli lipopolysaccharide (LPS) increased the number of alveolar type II cells in the fetal mouse lung. LPS also increased type II cell number in cultured fetal lung explants, suggesting that LPS could directly signal the fetal lung in the absence of maternal influences. Using immunostaining, we localized cells within the fetal mouse lung expressing the LPS receptor molecule Toll-like receptor 4 (TLR4). Similar to the signaling pathways in inflammatory cells, LPS activated NF-kappaB in fetal lung explants. Activation of the TLR4/NF-kappaB pathway appeared to be required, as LPS did not increase the number of type II cells in C.C3H-Tlr4(Lps-d) mice, a congenic strain containing a loss of function mutation in tlr4. In addition, the sesquiterpene lactone parthenolide inhibited NF-kappaB activation following LPS exposure and blocked the LPS-induced increase in type II cells. On the basis of these data from our mouse model of chorioamnionitis, it appears that LPS specifically activated the TLR4/NF-kappaB pathway, leading to increased type II cell maturation. These data implicate an important signaling mechanism in chorioamnionitis and suggest the TLR4/NF-kappaB pathway can influence lung development.     

Cytidine monophosphate-dependent synthesis of phosphatidylglycerol in permeabilized type II pneumonocytes.

Results of previous investigations support the proposition that, in type II pneumonocytes, CMP is involved in integration of the synthesis of phosphatidylcholine and phosphatidylglycerol for lung surfactant. In the present investigation, the amount of CMP in rat type II pneumonocytes was altered directly and resultant changes in the synthesis of phosphatidylglycerol were examined. Type II pneumonocytes were made permeable to CMP by treatment with Ca2+-free medium, and phosphatidylglycerol synthesis was then assessed by measurement of the incorporation of a radiolabelled precursor, [14C]glycerol 3-phosphate, that was not effectively utilized by cells that resisted permeabilization. Incorporation of [14C]glycerol 3-phosphate into phosphatidylglycerol (but not into other lipids) was stimulated greatly by CMP (half-maximal stimulation at approx. 0.1 mM). CMP stimulated the incorporation of [14C]glycerol 3-phosphate into both the phosphatidyl moiety and the head group of phosphatidylglycerol. Incorporation of [14C]palmitate into phosphatidylglycerol was also stimulated by CMP. myo-Inositol, at concentrations found in foetal-rat serum (0.2-2.0 mM), inhibited CMP-dependent incorporation of [14C]glycerol 3-phosphate into phosphatidylglycerol and promoted, instead, CMP-dependent incorporation into phosphatidylinositol. These data, when extrapolated to foetal type II pneumonocytes, are consistent with the view that the developmental increase in the synthesis of phosphatidylglycerol for surfactant by foetal lungs is promoted by the increase in intracellular CMP and the declining availability of myo-inositol that were found previously to be associated with this period of development.     

Sexually dimorphic serotonergic dysfunction in a mouse model of Huntington's disease and depression

Depression is the most common psychiatric disorder in Huntington's disease (HD) patients. In the general population, women are more prone to develop depression and such susceptibility might be related to serotonergic dysregulation. There is yet to be a study of sexual dimorphism in the development and presentation of depression in HD patients. We investigated whether 8-week-old male and female R6/1 transgenic HD mice display depressive-like endophenotypes associated with serotonergic impairments. We also studied the behavioral effects of acute treatment with sertraline. We found that only female HD mice exhibited a decreased preference for saccharin as well as impaired emotionality-related behaviors when assessed on the novelty-suppressed feeding test (NSFT) and the forced-swimming test (FST). The exaggerated immobility time displayed by female HD in the FST was reduced by acute administration of sertraline. We also report an increased response to the 5-HT(1A) receptor agonist 8-OH-DPAT in inducing hypothermia and a decreased 5-HT(2A) receptor function in HD animals. While tissue levels of serotonin were reduced in both male and female HD mice, we found that serotonin concentration and hydroxylase-2 (TPH2) mRNA levels were higher in the hippocampus of males compared to female animals. Finally, the antidepressant-like effects of sertraline in the FST were blunted in male HD animals. This study reveals sex-specific depressive-related behaviors during an early stage of HD prior to any cognitive and motor deficits. Our data suggest a crucial role for disrupted serotonin signaling in mediating the sexually dimorphic depression-like phenotype in HD mice.     

Regulation of CTP: phosphocholine cytidylyltransferase activity in type II pneumonocytes.

Phosphatidylcholine synthesis by rat type II pneumonocytes was altered either by depleting the cells of choline or by exposing the cells to extracellular lung surfactant. Effects of these experimental treatments on the activity of a regulatory enzyme, CTP:phosphocholine cytidylyltransferase, were investigated. Although choline depletion of type II pneumonocytes resulted in inhibition of phosphatidylcholine synthesis, cytidylyltransferase activity (measured in cell homogenates in either the absence or presence of added lipids) was greatly increased. Activation of cytidylyltransferase in choline-depleted cells was rapid and specific, and was quickly and completely reversed when choline-depleted cells were exposed to choline (but not ethanolamine). Choline-dependent changes in enzymic activity were apparently not a result of direct actions of choline on cytidylyltransferase and they were largely unaffected by cyclic AMP analogues, oleic acid, linoleic acid or cycloheximide. The Km value of cytidylyltransferase for CTP (but not phosphocholine) was lower in choline-depleted cells than in choline-repleted cells. Subcellular redistribution of cytidylyltransferase also was associated with activation of the enzyme in choline-depleted cells. When measured in the presence of added lipids, 66.5 +/- 5.0% of recovered cytidylyltransferase activity was particulate in choline-depleted cells but only 34.1 +/- 4.5% was particulate in choline-repleted cells. An increase in particulate cytidylyltransferase also occurred in type II pneumonocytes that were exposed to extracellular surfactant. This latter subcellular redistribution, however, was not accompanied by a change in cytidylyltransferase activity even though incorporation of [3H]choline into phosphatidylcholine was inhibited by approx. 50%. Subcellular redistribution of cytidylyltransferase, therefore, is associated with changes in enzymic activity under some conditions, but can also occur without a resultant alteration in enzymic activity.