[Discussion of grading method of small opacity profusion of pneumoconiosis on CT scans and the corresponding reference images].

Objective: To explore the CT grading method of small opacity profusion of pneumoconiosis, and draw up the corresponding CT reference film. Methods: In December 2019, Three hundred thirty-seven cases of pneumoconosis and suspected pneumoconiosis were examined by chest radiography and Computed Tomography (CT) in the same period. According to Diagnosis of Occupational Pneumoconiosis (GBZ 70-2015) , small opacity profusion of pneumoconiosis in each zone of lung was divided. On CT scans, it was divided into 5 grades of 0, 0+, 1, 2 and 3. Grade 0 corresponded to Sub-grade 0/- and Sub-grade 0/0 of Grade 0 in chest radiograph. Grade 0+ was equivalent to Sub-grade 0/1 of Grade 0. Grade 1, 2, 3 were equivalent to Grade 1, 2 and 3, respectively (including each sub-grade) . The CT image quality of each zone of lung was divided into 1 to 4 levels. Results of level 4 were not included in statistical analyses.Based on the results of small opacity profusion in each zone of lung, consistency analysis was performed between chest radiograph and CT. The selection method of reference films was developed. Based on the types and grades of small opacity, the final reference films were determined. Results: There were 1877 zones of lung with CT image quality from level 1 to 3, including 335 in upper right, 319 in middle right, 284 in lower right, 334 in upper left, 320 in middle left and 286 in lower left. The Kappa values of small opacity profusion in upper right zone, upper left zone, left middle zone, and lower left zone were all between 0.4-0.75. In middle right zone and lower right zone, they were all above 0.75.Among all 6 zones of lung, the diagnostic concordance rates between CT and chest radiograph were all above 80%.The corresponding CT reference films were proposed, including type p and q in Grade 2 and 3, type r in Grade 2, type s and t in Grade 0+ to 3. Conclusion: The CT grading method for small opacity profusion of pneumoconiosis is feasible, and the application value of its reference films needs to be further verified.

Analysis of imprinted messenger RNA expression in deceased transgenic cloned goats.

Genomic imprinting is an important epigenetic mechanism that has vital effects on fetal growth and development. We observed the differences in four tissues (heart, spleen, liver, and kidney) from dead transgenic cloned goats using hematoxylin and eosin (H&E) staining. Eight imprinted genes in the tissues of dead transgenic cloned and normal goats were analyzed using reverse transcription polymerase chain reaction. H&E staining results from the abortion group indicated the lack of obvious morphological changes in heart and spleen tissues, while inflammatory cell infiltration and glomerular nephritis characteristics were observed in liver and kidney tissues, respectively. Compared to the control group, CDKN1C, H19, IGF2R, and SNRPN were significantly (P < 0.05) overexpressed in the heart tissue of the abortion group, while XIST was significantly reduced. In the liver tissues, CDKN1C and DLK1 expression decreased, while GNAS, H19, IGF2R, PEG3, and XIST expression increased significantly. In the spleen tissues, DLK1 expression increased, while GNAS, H19, IGF2R, PEG3, SNRPN, and XIST expression decreased. In the kidney tissues, CDKN1C, DLK1, GNAS, IGF2R, and PEG3 expression increased, while H19 and XIST expression decreased. The overall expression of imprinted genes was abnormal in different tissues of transgenic cloned goats, and the degree of abnormal genomic imprinting was more severe in the abortion group compared to the death and control groups. These results suggest that abnormal expression of imprinted genes may cause developmental defects in transgenic cloned goats. Moreover, abnormal epigenetic modifications may affect the reprogramming of transgenic donor cells.

Hemodynamic changes of unexplained syncope patients in head-up tilt test.

The purpose of this study was to observe the hemodynamic changes of unexplained syncope patients in the head-up tilt test and their correlations with age and gender. Eighty-six patients with unexplained syncope were administered the basic head-up test and nitroglycerin provocation test with continuous monitoring and recording of electrocardiogram and blood pressure changes. Basic characteristics of the patients and their hemodynamic responses throughout the tests were analyzed. All 86 patients tolerated and completed the head-up test. Forty-nine (56.98%) of the patients displayed a positive reaction, 37 (43.02%) patients displayed a negative reaction. Patients were divided into groups as follows: Group A, age ≤ 35 years; Group B, age 36-45 years; and Group C, age ≥ 46 years. Older patients were more prone to chronotropic incompetence, and younger patients were more prone to an excessive increase in heart rate. Older age correlated with the occurrence of autonomic nerve reaction disorder and mixed vasovagal syncope, whereas younger age was related to the occurrence of vasodepressor type vasovagal syncope (P < 0.01). Gender did not significantly correlate with negative or positive head-up test results (P = 0.184). During the head-up test, younger patients mainly manifested an excessive heart rate increase, whereas older patients did not have significant heart rate changes. Analyzing the hemodynamic changes in the head-up test and studying the relationships between age, gender, and hemodynamic responses are crucial to determine etiologies of syncope and select appropriate treatment.

Identification of transgenic cloned dairy goats harboring human lactoferrin and methylation status of the imprinted gene IGF2R in their lungs.

Dairy goat is a good model for production of transgenic proteins in milk using somatic cell nuclear transfer (SCNT). However, animals produced from SCNT are often associated with lung deficiencies. We recently produced six transgenic cloned dairy goats harboring the human lactoferrin gene, including three live transgenic clones and three deceased transgenic clones that died from respiratory failure during the perinatal period. Imprinted genes are important regulators of lung growth, and may be subjected to faulty reprogramming. In the present study, first, microsatellite analysis, PCR, and DNA sequence identification were conducted to confirm that these three dead kids were genetically identical to the transgenic donor cells. Second, the CpG island methylation profile of the imprinted insulin-like growth factor receptor (IGF2R) gene was assessed in the lungs of the three dead transgenic kids and the normally produced kids using bisulfite sequencing PCR. In addition, the relative mRNA level of IGF2R was also determined by real-time PCR. Results showed that the IGF2R gene in the lungs of the dead cloned kids showed abnormal hypermethylation and higher mRNA expression levels than the control, indicating that aberrant DNA methylation reprogramming is one of the important factors in the death of transgenic cloned animals.

Effect of Age on Energy Requirement for Maintenance and Growth of Dorper and Hu Crossbred F1 Ewes Weighing 20 to 50 kg.

This research aimed to define the energy requirement of Dorper and Hu Hybrid F1 ewes 20 to 50 kg of body weight, furthermore to study energy requirement changes with age and evaluate the effect of age on energy requirement parameters. In comparative slaughter trial, thirty animals were divided into three dry matter intake treatments (ad libitum, n = 18; low restricted, n = 6; high restricted, n = 6), and were all slaughtered as baseline, intermediate, and final slaughter groups, to calculate body chemical components and energy retained. In digestibility trial, twelve ewes were housed in individual metabolic cages and randomly assigned to three feeding treatments in accordance with the design of a comparative slaughter trial, to evaluate dietary energetic values at different feed intake levels. The combined data indicated that, with increasing age, the net energy requirement for maintenance (NEm) decreased from 260.62±13.21 to 250.61±11.79 kJ/kg(0.75) of shrunk body weight (SBW)/d, and metabolizable energy requirement for maintenance (MEm) decreased from 401.99±20.31 to 371.23±17.47 kJ/kg(0.75) of SBW/d. Partial efficiency of ME utilization for maintenance (km, 0.65 vs 0.68) and growth (kg, 0.42 vs 0.41) did not differ (p>0.05) due to age; At the similar condition of average daily gain, net energy requirements for growth (NEg) and metabolizable energy requirements for growth (MEg) for ewes during late fattening period were 23% and 25% greater than corresponding values of ewes during early fattening period. In conclusion, the effect of age upon energy requirement parameters in the present study were similar in tendency with previous recommendations, values of energy requirement for growth (NEg and MEg) for Dorper and Hu crossbred female lambs ranged between the NRC (2007) recommendation for early and later maturating growing sheep.

Hepatic DNA methylation modifications in early development of rats resulting from perinatal BPA exposure contribute to insulin resistance in adulthood.

AIMS/HYPOTHESIS: Perinatal exposure to bisphenol A (BPA), a widely distributed environmental endocrine disruptor, is associated with insulin resistance and diabetes in offspring. The underlying molecular mechanisms could involve epigenetics, as adverse effects induced by environmental exposure in early life are suggested through DNA methylation. In this study we sought to elucidate the relationship between perinatal BPA exposure and alteration of hepatic DNA methylation. METHODS: Pregnant Wistar rats were administered BPA (50 µg/kg/day) or corn oil by oral gavage throughout gestation and lactation. Variables associated with insulin resistance and hepatic DNA methylation were examined at postnatal week 3 and week 21 in male offspring. RESULTS: In BPA-treated offspring, serum insulin and HOMA-insulin resistance were increased, and the insulin sensitivity index and hepatic glycogen storage were decreased compared with controls at week 21. At week 3, none of these variables were significantly changed. However, hepatic global DNA methylation was decreased, accompanied by overexpression of DNA methyltransferase 3B mRNA at week 3. Meanwhile, perinatal exposure to BPA induced promoter hypermethylation and a reduction in gene expression of hepatic glucokinase. Moreover, increased promoter hypermethylation of Gck became more pronounced in BPA-treated offspring at week 21. CONCLUSIONS/INTERPRETATION: Abnormal DNA methylation in hepatic tissue precedes development of insulin resistance induced by perinatal BPA exposure. These findings support the potential role of epigenetics in fetal reprogramming by BPA-induced metabolic disorders.

Monoclonal antibodies to laminin reveal the heterogeneity of basement membranes in the developing and adult mouse tissues.

Two monoclonal antibodies raised against laminin isolated from a mouse parietal yolk sac cell line were used for immunohistochemical studies of basement membranes of the mouse embryo and various fetal and adult tissues. No immunoreactivity with either of the two monoclonal antibodies could be detected in the preimplantation-stage embryos, although it has been shown that these embryos contain extracellular laminin reactive with the conventional polyclonal antilaminin antibodies. Reichert's membrane in early postimplantation stages of development reacted with the monoclonal antibody LAM-I but not with the antibody LAM-II. However, from day 8 of pregnancy onward the Reichert's membrane reacted with both antibodies. Basement membranes of the embryo proper were unreactive with both monoclonal antibodies until day 12 of pregnancy. By day 14 some basement membranes of the fetal tissues became reactive with one or both monoclonal antibodies, whereas others remained still unreactive. In the 17-d fetus and the newborn mouse most of the basement membranes reacted with both monoclonal antibodies, whereas others still reacted with only one. Similar heterogeneity in the immunoreactivity of basement membranes of various tissues was noted in the adult mouse as well. These results indicate that the immunoreactivity of laminin in the extracellular matrix changes during development and that the basement membranes in various anatomic locations display heterogeneity even in the adult mouse.

Edaravone protects from retinal injury through NF-κB in diabetic rats.

OBJECTIVE: To observe whether edaravone has a therapeutic and protective effect on retinal injury in diabetic rats through the nuclear factor-κB (NF-κB) pathway. MATERIALS AND METHODS: The Sprague-Dawley rat model of diabetes was established and divided into diabetes model group (model group) and edaravone treatment group (treatment group). Also, normal control group (control group) was set up. After successful modeling, the blood and retinal tissues of rats were collected. Then, the blood glucose content and serum interleukin-6 (IL-6) were detected. Antioxidant indexes, superoxide dismutase (SOD), and malondialdehyde (MDA), were detected via enzyme-linked immunosorbent assay (ELISA), while the number of corneal nerve fibers was observed microscopically. Moreover, the gene and protein expressions of SOD and NF-κB pathway in tissues were detected via quantitative Reverse Transcription-Polymerase Chain Reaction (qRT-PCR) and Western blotting. RESULTS: The level of blood glucose in model group was increased compared with that in control group (p<0.05), indicating the successful modeling. The levels of tumor necrosis factor-α (TNF-α), IL-6, and IL-1 were significantly higher in model group than those in control group. In terms of the antioxidant indexes, the level of MDA in model group was significantly higher than that in other two groups, while the level of SOD in treatment group was significantly increased and close to that in control group. Besides, the number of corneal nerve fibers significantly declined in model group, while it was increased in treatment group but still lower than that in control group. According to the gene detection results, the mRNA expression of NF-κB p65 was markedly higher in model group than that in control group, and it was decreased in treatment group, while the mRNA expression of SOD showed the opposite trend. The protein expression of NF-κB p65 was remarkably higher in model group than in control group, was decreased in treatment group, and was close to that in control group, while the protein expression of SOD showed the opposite trend. CONCLUSIONS: Edaravone can affect the oxidation and antioxidation through the NF-κB signaling pathway, thereby exerting a therapeutic and protective effect on retinal injury in diabetic rats.

A burst of c-fos gene expression in the mouse occurs at birth.

Expression of the c-fos gene during murine perinatal development was studied. Before birth, all eight of the prenatal organs tested expressed undetectable or low levels of c-fos mRNA. On the day of birth, there occurred a 10- to 100-fold increase in the level of c-fos message in all of these organs. The expression was transient, in that 1 day after birth, the level of c-fos mRNA precipitously dropped. The c-fos gene expression at birth is unrelated to the expression of the c-myc gene and major histocompatibility complex class I genes, which display distinct kinetics during the perinatal development. The c-fos gene was also expressed locally and transiently in the gravid uterus 1 to 2 days prior to delivery. These results indicate that an event associated with birth induced c-fos gene expression in the mother and newborn.

Regulation of rat liver maturation in vitro by glucocorticoids.

The biochemistry of liver maturation was studied by using the RLA209-15 fetal rat hepatocyte line that is temperature sensitive for maintenance of the differentiated fetal liver phenotype. At 33 degrees C these cells were dedifferentiated; but at 40 degrees C they were phenotypically differentiated and, like normal fetal hepatocytes, synthesized moderate levels of albumin and transferrin, high levels of authentic (69,000 and 73,000 molecular weight) rat fetal alpha-fetoprotein (AFP), and low levels of a 65,000-molecular-weight variant AFP. Our results indicated that administration of glucocorticoid hormones to RLA209-15 cells at 40 degrees C induced a series of events associated with normal hepatocyte maturation; synthesis of fetal AFP was inhibited, whereas the synthesis of variant AFP, albumin, transferrin, tyrosine aminotransferase, and alpha 1-acid glycoprotein was induced. The variant AFP was produced by RLA209-15 cells at both temperatures and was encoded by an mRNA of 1.7 kilobases (kb). The fetal AFP was encoded by an mRNA of 2.2 kb. Normal adult rat liver contained three AFP mRNAs of 2.2 (minor), 1.7, and 1.5 kb. The 1.7-kb adult liver AFP mRNA comigrated with the RNA found in RLA209-15 cells, and both directed the synthesis of a 50,000-molecular-weight precursor polypeptide of the variant AFP. Administration of glucocorticoids to RLA209-15 cells grown at 33 degrees C stimulated synthesis of both the fetal and variant AFPs, but the levels of the 2.2-kb AFP mRNA were preferentially increased. RLA209-15 cells contained two glucocorticoid receptor mRNAs of 6.8 and 4.5 kb. The glucocorticoid-mediated maturation described above was blocked by the antiglucocorticoid RU486.

Hepatocyte-specific mutation establishes retinoid X receptor alpha as a heterodimeric integrator of multiple physiological processes in the liver.

A large number of physiological processes in the adult liver are regulated by nuclear receptors that require heterodimerization with retinoid X receptors (RXRs). In this study, we have used cre-mediated recombination to disrupt the mouse RXRalpha gene specifically in hepatocytes. Although such mice are viable, molecular and biochemical parameters indicate that every one of the examined metabolic pathways in the liver (mediated by RXR heterodimerization with PPARalpha, CARbeta, PXR, LXR, and FXR) is compromised in the absence of RXRalpha. These data demonstrate the presence of a complex circuitry in which RXRalpha is integrated into a number of diverse physiological pathways as a common regulatory component of cholesterol, fatty acid, bile acid, steroid, and xenobiotic metabolism and homeostasis.

Application of levator veli palatini retropositioning combined with Buccinator myomucosal island flap for congenital cleft palate.

Congenital cleft palate causes a serious obstacle to children with regard to language and eating function. The aim of the current study was to examine the clinical application of a type of palatoplasty that has a reduced impact on the maxillary growth and good function in velopharyngeal competence. A total of 37 patients with cleft palate were treated with levator veli palatini retropositioning combined with Buccinator myomucosal island flap. The patients were successfully treated in the first phase and were followed up for 1-3 years. Speech intelligibility was satisfactory and no fistula occurred. In conclusion, the results suggested that levator veli palatini retropositioning combined with the Buccinator myomucosal island flap may restore normal anatomic structure and location of the levator veli palatini, obtain good velopharyngeal competence, and decrease the incidence rate thereof. Thus, levator veli palatini retropositioning combined with the Buccinator myomucosal island flap is a functional procedure for cleft palate repair.

Characterization of pregnancy-specific beta 1-glycoprotein synthesized by human placental fibroblasts.

We have previously demonstrated that human placental fibroblasts produce a pregnancy-specific beta 1-glycoprotein (PS beta G) immunologically indistinguishable from placental PS beta G. This was confirmed by the immunocytochemical localization of PS beta G in these fibroblasts. In addition, placental fibroblasts contain all three PS beta G mRNAs of 2.3, 2.2, and 1.7 kilobases which hybridize with the three PS beta G cDNAs (PSG16, PSG93, and PSG95) identified, although at 1.4-2.5% of the levels in human term placenta. The major PS beta G species synthesized by placental fibroblasts is a 62K glycopolypeptide formed from a 58K intracellular precursor polypeptide. However, the PS beta G species found in human placenta are one major glycoprotein of 72K and two minor ones of 64K and 54K. Poly(A)+ RNA from placental fibroblasts directed the synthesis of two polypeptides of 48K and 46K (major), whereas, poly(A)+ RNA from human placenta directed the synthesis of higher levels of four polypeptides of 50 K, 48 K (major), 46 K, and 36 K. Thus, the major PS beta G species found in fibroblasts and human placenta differ. The carbohydrate side-chains are essential for the stability of fibroblast PS beta G, because PS beta G synthesis in these fibroblasts could not be detected in the presence of tunicamycin, a protein glycosylation inhibitor which did not affect PS beta G mRNA expression. Our finding that a variant PS beta G species is produced in placental fibroblasts raises the possibility that the authentic placental PS beta G species may have different functions.

Inhibin subunit gene expression and distribution in the ovaries of immature, young adult, middle-aged, and old female rats.

In young adult female rats, the patterns of inhibin subunit mRNA expression during the estrous cycle are regulated by cyclic changes in gonadotropin secretion and follicular development. Since there are distinct alterations in profiles of both hormone secretion and folliculogenesis during the reproductive lifespan of the female rat, we have characterized the gene expression and distribution of inhibin subunit mRNAs in immature (22 days), young adult cyclic (3-4 months), middle-aged cyclic (9-10 months), and old (12-13 months) persistent estrous (PE) rat ovaries. Northern blot analyses revealed that in contrast to young adult cyclic rats, inhibin alpha- and beta A-subunit mRNA levels in the ovaries of middle-aged cyclic rats remained substantially elevated after the proestrous gonadotropin surges and ovulation. Likewise, acyclic immature and old PE rats showed high levels of inhibin alpha- and beta A-subunit transcripts in their ovaries. In situ hybridization analyses demonstrated that inhibin alpha- and beta A mRNAs were abundantly expressed in maturing follicles of both young and middle-aged cyclic females, while high levels of alpha-subunit transcripts were only detected in the ovarian stroma of middle-aged animals. The ovaries of old PE rats had numerous large cystic follicles (with variable layers of granulosa cells) and few degenerating cyst-like structures (completely devoid of granulosa cells). Inhibin subunit transcripts were expressed abundantly in both the granulosa (alpha and beta A-subunits) and theca interna (alpha-subunit only) layers of large follicles, but were absent from degenerating cysts devoid of granulosa cells. The ovarian stroma of PE rats also expressed very high levels of inhibin-alpha, but not beta A mRNA. The ovaries of immature rats contained large numbers of uniformly developing secondary follicles. High levels of inhibin-alpha mRNA were expressed homogeneously in the granulosa layer of all growing follicles, whereas inhibin beta A mRNAs were only detected in selectively larger follicles with multiple layers of granulosa cells. Hormone RIAs of serum samples from these same groups of animals showed that basal levels of serum FSH were substantially higher in immature, middle-aged cyclic, and old PE rats than in young adult rats. These results demonstrate that enhanced ovarian inhibin subunit gene expression in the female rat is associated with increased serum FSH levels regardless of chronological age. On the other hand, aging appears to selectively enhance inhibin alpha, but not beta A, gene expression in the ovarian stroma, such that it may gradually become a major secondary site of alpha-subunit mRNA production in addition to the follicular compartments.

Analysis of the CYP2D6 gene polymorphism and enzyme activity in African-Americans in southern California.

Despite its importance in drug metabolism and disease susceptibility, CYP2D6 activity and genetic polymorphism have rarely been investigated in African-American populations. In order to bridge this gap, we examined the genotype and phenotype of the enzyme in 154 African-American (AA) and 143 Caucasian (C) normal volunteers. AAs are significantly more likely to possess *17 and *5, but less likely to have *4. Overall, the two groups were similar in their CYP2D6 activity as measured with dextromethorphan as the probe (metabolic ratio 2.21 +/- 0.78 for AAs; 2.11 +/- 0.86 for Cs; t = 1.02, NS). Two of four AAs and six of seven Cs were classified as poor metabolizers and have two nonfunctioning alleles. CYP2D6 activity is determined by *17, *4, *5 and age in AAs (r2 = 0.33, f = 18.8, P < 0.001) and by *4 and *XN in Cs (r2 = 0.14, f = 10.8, P < 0.001). These results support previous findings demonstrating the importance of *17 in determining CYP2D6 activity in AAs.

CYP2D6 polymorphism in a Mexican American population.

BACKGROUND: Pharmacogenetic data are largely unavailable for Mexican Americans, despite being one of the largest populations in America. METHODS: The CYP2D6 genotype (n = 349) and dextromethorphan hydroxylation phenotype (n = 285) were studied in 380 Mexican American subjects from Los Angeles County. RESULTS: The allelic frequency was 22.8% for CYP2D6*2, 10.3% for CYP2D6*4, 7.4% for CYP2D6*10, 2.3% for CYP2D6*5, 1% for CYP2D6*XN (duplication), and <1% for CYP2D6*3 and CYP2D6*17. By using the published antimode for Caucasians, we identified nine subjects as poor metabolizers, an incidence of 3.2%. Of the eight poor metabolizers who were also genotyped, five either were homozygous for the CYP2D6*4 allele (4 cases) or had a combination of CYP2D6*4 and CYP2D6*5 alleles. The mean log(10) dextromethorphan/dextrorphan ratio was -2.47 for those classified as extensive metabolizers. The number of functional alleles among the extensive metabolizers correlated strongly with the phenotype, suggesting a gene-dose effect. CONCLUSION: Compared with previous reports on Caucasian populations, studies show that Mexican Americans appear to possess a lower rate of CYP2D6*4. Frequencies for the other alleles appear to be less divergent between the two groups. This genotypic pattern might be responsible for the lower rate for the poor metabolizer status, as well as for the faster enzyme activity in the extensive metabolizer subjects that was also reflected in our data.

Role of CYP2E1 in the pathogenesis of alcoholic liver disease: modifications by cAMP and ubiquitin-proteasome pathway.

The ethanol inducible isoform of cytochrome P450, CYP2E1, may play a role in ethanol-induced liver injury. Therefore, the factors which govern CYP2E1 degradation and turnover were investigated. These factors include cAMP, ubiquitin, proteasomal enzymes and CYP2E1 mRNA. Rats fed ethanol or pair-fed isocaloric dextrose were pair-fed with rats fed ethanol or dextrose treated with cAMP for 2 months. The liver pathology, regenerative activity, fatty acid composition, NFkappaB activation, ubiquitin conjugates and proteasomal enzymes were measured as were the apoprotein levels of CYP2E1, CYP3A, CYP4A and mRNA levels for CYP2E1 and ubiquitin expression. The results showed, that the cAMP treatment ameliorated the increase liver fat storage and changes in the fatty acid composition in the livers of ethanol fed rats. Other histologic features of alcoholic liver disease were not changed. Western blot quantitation showed that the amount of ubiquitin and ubiquitin conjugates were markedly reduced by ethanol treatment. Similarly, ethanol decreased the level of ubiquitin mRNA. cAMP ameliorated the inhibition of the proteasomal enzyme proteolysis caused by ethanol feeding. The ethanol-induced increase in the CYP2E1 protein was partially inhibited by cAMP treatment. cAMP treatment decreased CYP2E1 mRNA levels in both ethanol-fed and pair fed control rats. Likewise NFkappaB activation was not increased by ethanol but cAMP reduced the level of NFkappaB activation. CAMP treatment also reduced CYP4A but not CYP3A. The results support the concept that cAMP treatment partially protects the liver from ethanol-induced fatty liver by reducing CYP2E1 induction through cAMP's effects on CYP2E1 synthesis.

Detection of retinoic acid receptor mRNA in rat tissues by reverse transcriptase-polymerase chain reaction.

The presence of retinoic acid receptor (RAR) alpha, beta and gamma mRNA was examined in 16 different kinds of rat tissue using the highly sensitive reverse transcriptase-polymerase chain reaction technique. The data demonstrated that each tissue expressed at least two types of RAR mRNA. Among the three types of RAR mRNA, RAR alpha was widely expressed in all types of organ and was the dominant form expressed in the gastrointestinal tract. RAR beta mRNA was not present in the intestine and spleen. In addition, RAR beta mRNA levels were high in the heart, lung, brain, testis and epididymis. RAR gamma mRNA was abundant in both male and female reproductive systems, as well as epidermal tissues. The prevalence of each RAR mRNA in the tissues suggests the diverse biological roles of these receptors.

Retinoic acid induces activin receptor IIB mRNA in F9 embryonal carcinoma cells.

Mouse embryonal carcinoma F9 cells are pluripotent stem cells and differentiate into primitive endodermal cells upon treatment with retinoic acid (RA). We have recently shown that in F9 cells RA regulates gene expression of activin receptor type II (ActR-II), whose ligand is a potent differentiation agent. The present study examined the regulation of the newly cloned activin receptor type IIB (ActR-IIB) gene by RA. F9 cells expressed equal amounts of three ActR-IIB transcripts of 8.0, 7.5 and 4.0 kb. Both 9-cis-RA (c-RA) and all-trans-RA (t-RA) induced ActR-IIB gene expression in a dose-dependent manner. At 10(-9) M c-RA exerted no effect, while 10(-5) M c-RA increased the 8.0 kb ActR-IIB transcript about sevenfold. In contrast, t-RA induced the 8.0 kb ActR-IIB transcript fivefold at 10(-9) M and up to eightfold at 10(-5) M. The inductive effect on the 8.0 kb transcript was greater than that on the 7.5 kb transcript, and was least effective on the 4.0 kb transcript, suggesting that these three mRNA isoforms may originate from different promoters. Both cycloheximide and actinomycin D inhibited the inductive effect of t-RA on ActR-IIB gene expression, in contrast to ActR-II whose gene expression was not suppressed by cycloheximide but abolished by actinomycin D. Thus, endodermal differentiation of F9 cells is associated with activation of ActR-IIB gene and the mechanisms involved in the regulation of ActR-II and IIB gene expression are different.

9-cis-retinoic acid is more effective than all-trans-retinoic acid in upregulating expression of the alpha-fetoprotein gene.

In McA-RH 8994 rat hepatoma cells, all-trans-retinoic acid (t-RA) induces expression of the alpha-fetoprotein (AFP) and albumin genes and results in a phenotype similar to differentiated fetal hepatocytes. The present study elucidated the mechanism involved in AFP gene regulation mediated by retinoic acid. Northern blot analyses demonstrated that 9-cis-retinoic acid (c-RA), a ligand for retinoid x receptors (RXRs), also induced expression of the AFP gene in McA-RH 8994 cells. The induction was time- and dose-dependent. Northern blots and transfection assays using the 7.3 kb full-length regulatory region of the AFP gene demonstrated that c-RA was more effective than t-RA in regulating expression of the AFP gene. At 10(-7) M, c-RA increased AFP mRNA 5-fold and chloramphenicol acetyltransferase (CAT) activity 2.5-fold. In contrast, t-RA at a concentration of 10(-7) M exerted no significant effect; 10(-6) to 10(-5) M t-RA was needed to affect AFP gene expression. These data suggested that activation of RXRs is essential for the regulation of the AFP gene. Co-transfection experiments revealed that over-expression of RXR alpha in McA-RH 8994 cells further enhanced the CAT activity induced by c-RA. In addition, c-RA did not alter the half-life of AFP mRNA. Thus, RXR alpha may play a crucial role in transcriptional regulation of the AFP gene and in controlling hepatocyte phenotype.