Cathepsin D mediates prenatal caffeine exposure-caused NAFLD susceptibility in male rat offspring by regulating autophagy.

Epidemiological evidence has revealed that non-alcoholic fatty liver disease (NAFLD) harbors an intrauterine origin. Autophagy is known to be involved in the protective mechanism in the development of adult NAFLD, but whether it engages in the occurrence of fetal-originated NAFLD remains unclear. In this study, a rat model of fetal-originated NAFLD was established by giving a high-fat diet or chronic stress after birth on prenatal caffeine exposure (PCE) male offspring. The alterations of liver morphologic analysis, lipid metabolism, and autophagy before and after birth were determined to confirm autophagy mechanism, NAFLD susceptibility, and intrauterine origin in PCE male adult offspring. Our results revealed that PCE-induced intrauterine high concentration of corticosterone exposure blocked autophagic flux by inhibiting cathepsin D expression in hepatocytes, leading to ß-oxidation inhibition and lipid accumulation in the liver. Moreover, high concentration of corticosterone upregulated miR-665 by activating the glucocorticoid receptor to suppress cathepsin D, thus causing lysosomal degradation dysfunction and autophagy flux blockade. Notably, hepatic overexpression of cathepsin D could reverse PCE-induced postnatal NAFLD susceptibility in male rat offspring. This study elucidates the epigenetic programming mechanism of intrauterine autophagy-related fetal-originated NAFLD susceptibility, and identifies cathepsin D as its early intervention target, providing an experimental basis for exploring early prevention and treatment strategies for fetal-originated NAFLD.

[Factors Influencing and Adverse Reactions of Voriconazole Clearance in Patients with Hematological Diseases].

OBJECTIVE: To monitor the changes of voriconazole minimum concentration(Cmin) in patients with hematological diseases, and evaluate the factors influencing and adverse reactions of voriconazole clearance in patients with hematological diseases, so as to provide a theoretical basis for reasonable clinical use of voriconazole. METHODS: 136 patients with hematological diseases who used voriconazole in Wuhan NO.1 Hospital from May 2018 to December 2019 were selected. The correlation between C-reactive protein, albumin, creatinine and voriconazole Cmin were analyzed, and the changes of voriconazole Cmin after glucocorticoid treatment was also detected. In addition, stratified analysis was used to explore the adverse events of voriconazole. RESULTS: Among 136 patients, 77 were male (56.62%) and 59 were female (43.38%). There were positive correlations between voriconazole Cmin and C-reactive protein and creatinine levels (r=0.277, r=0.208), while voriconazole Cmin was negatively correlated with albumin level (r=-2.673). Voriconazole Cmin in patients treated with glucocorticoid was decreased significantly (P<0.05). In addition, sratified analysis of voriconazole Cmin showed that compared with voriconazole Cmin 1.0-5.0 mg/L group, the incidence of adverse reactions of visual impairment in voriconazole Cmin> 5.0 mg/L group was increased (χ2=4.318, P=0.038). CONCLUSION: The levels of C-reactive protein, albumin and creatinine are closely related to the voriconazole Cmin, which indicate that inflammation and hyponutrition may prevent the clearance of voriconazole in patients with hematological diseases. It is necessary to monitor the voriconazole Cmin of patients with hematological diseases, and adjust the dosage in time to reduce adverse reactions.

Sex differences and heritability of adrenal steroidogenesis in offspring rats induced by prenatal nicotine exposure.

The epidemiological investigation has suggested prenatal nicotine exposure (PNE) induces multiorgan developmental toxicity and increases the risk of metabolic diseases in offspring. Our previous study found that the occurrence of fetal-originated diseases was associated with abnormal adrenal development in offspring. However, the long-term harmful effects on adrenal development in offspring induced by PNE remain unclear. Pregnant Wistar rats were injected subcutaneously with nicotine (2 mg/kg·d) from gestation day (GD) 9 to GD20 to obtain the adrenal gland from fetal and adult offspring rats of F1 and F2 generations. We found that the adrenal insulin-like growth factor 1 (IGF1) signaling pathway and steroidogenic function were increased in male while decreased in female of PNE fetal rats, which could extend into adulthood. Furthermore, the primary adrenal cells of fetal rats were treated with nicotine to observe the phenomena and clarify the possible mechanism of the sex difference. The results suggested that there are sex differences in IGF1 signaling pathway and steroidogenic function induced by PNE, which may be associated with sex differences in nAChRß1 expression. In addition, the adrenal steroidogenic function was reduced in F2 offspring of F1 PNE female rats (regardless of mating with control or Male PNE rats). Therefore, the decrease of adrenal steroidogenic function in female offspring rats induced by PNE has maternal heritability. In conclusion, PNE could lead to sex differences and heritability of adrenal steroidogenic function in offspring rats.

Prenatal dexamethasone exposure caused fetal rats liver dysplasia by inhibiting autophagy-mediated cell proliferation.

As a synthetic glucocorticoid, dexamethasone has been widely used in the clinical treatment of premature birth and related pregnant diseases, but its clinical use is still controversial due to developmental toxicity. This study aimed to confirm the proliferation inhibitory effect of pregnant dexamethasone exposure (PDE) on fetal liver development and elucidate its molecular mechanism. In vitro studies, we found that dexamethasone inhibited hepatocyte proliferation through autophagy activated by glucocorticoid receptor (GR)-forkhead protein O1 (FOXO1) pathway. Subsequently, in vivo, we confirmed in a PDE rat model that male fetal liver proliferation was inhibited, and the expression of the GR-FOXO1 pathway and autophagy were increased. Taken together, PDE induces autophagy by activating the GR-FOXO1 pathway, which leads to fetal liver proliferation inhibition and dysplasia in offspring rats. This study confirmed that dexamethasone activates cell autophagy in utero through the GR-FOXO1 pathway, thereby inhibiting hepatocyte proliferation and liver development, which provides theoretical basis for understanding the developmental toxicity of dexamethasone and guiding the rational clinical use.

Glucocorticoid-activation system mediated glucocorticoid-insulin-like growth factor 1 (GC-IGF1) axis programming alteration of adrenal dysfunction induced by prenatal caffeine exposure.

Glucocorticoids play a major factor in fetal maturation and fate decision after birth. We have previously demonstrated that prenatal caffeine exposure (PCE) resulted in adrenal dysplasia. However, its molecular mechanism has not been clarified. In the present study, a rat model of intrauterine growth retardation (IUGR) was established by PCE, and offspring were sacrificed. Moreover, NCI-H295 A cells were used to confirm glucocorticoid-related molecular mechanism. Results showed that PCE fetal weight decreased, and the IUGR rate increased, while serum corticosterone levels increased but insulin-like growth factor 1 (IGF1) levels decreased. Fetal adrenals exhibited an activated glucocorticoid-activation system, and the downregulated expression of IGF1 signal pathway and steroidal synthetases. For adult rats, there was no significant change in the glucocorticoid-activation system in the PCE group, the IGF1 signal pathway showed increased trend, and the expression levels of adrenal steroidal synthetases were close to normal. The data in vitro showed that the cortisol of 1200 nM can inhibit the expression of adrenocortical cell steroidal synthetases and IGF1 signal pathway when compared with the control. Meanwhile, the glucocorticoid-activation system was activated while GR inhibitor mifepristone can reverse the effect of cortisol. Furthermore, cortisol can also promote GR into the nucleus after its activation. Based on these findings, we speculated that high concentrations of glucocorticoid in utero led to GR in the nucleus through its activation and then inhibited the IGF1 signaling pathway by activating the glucocorticoid-activation system, which could further downregulate steroid synthesis.

Autophagy as a compensation mechanism participates in ethanol-induced fetal adrenal dysfunction in female rats.

Autophagy plays a vital role in embryonic development and cell differentiation. Our previous study demonstrated that prenatal ethanol exposure (PEE) resulted in intrauterine growth retardation (IUGR) and adrenal developmental toxicities in rat offspring. The present study focused on PEE-induced autophagy as an underlying mechanism and its biological significance in female fetal rats. Female fetuses in the PEE group exhibited lower body weights and suffered adrenal structural abnormalities compared to the controls. Cell proliferation was inhibited, the insulin-like growth factor 1 (IGF1) pathway was reduced, and autophagy was activated in the glands of female fetal rats. Ethanol increased the ratio of microtubule-associated protein light chain 3 beta-II (LC3ß-II) to LC3ß-I in vitro, and it reduced cortisol levels in time- and concentration-dependent manners in human adrenocortical carcinoma cells (NCI-H295A). Bafilomycin A1 inhibited autophagy, steroidogenic factor 1 (SF1) protein and steroidogenesis in the present study. Rapamycin with ethanol up-regulated autophagy and SF1 expression and activated steroidogenesis when compared with ethanol alone. In addition, ethanol inhibited IGF1 receptor (IGF1R) and phospho-mTOR (Ser2448) expression in a concentration-dependent manner. These results demonstrate that PEE activated autophagy in fetal adrenal glands, and the underlying mechanism may be associated with inhibition of the IGF1R/phospho-mTOR (Ser2448) pathway. Autophagy may be a compensatory mechanism for the PEE-induced inhibition of fetal adrenal steroidogenesis to maintain fetal adrenal development.

IGF1/MAPK/ERK signaling pathway-mediated programming alterations of adrenal cortex cell proliferation by prenatal caffeine exposure in male offspring rats.

Our previous study proposed a glucocorticoid-insulin-like growth factor 1 (GC-IGF1) axis programming mechanism for prenatal caffeine exposure (PCE)-induced adrenal developmental dysfunction. Here, we focused on PCE-induced cell proliferation changes of the adrenal cortex in male offspring rats before and after birth and clarified the intrauterine programming mechanism. On gestational day (GD) 20, the PCE group had an elevated serum corticosterone level reduced fetal bodyweight, maximum adrenal sectional area, and elevated adrenal corticosterone and aldosterone contents. However, in postnatal week (PW) 6, the serum corticosterone level was decreased, and the bodyweight, with catch-up growth, adrenal cortex maximum cross-sectional area and aldosterone content were relatively increased, while the adrenal corticosterone content was lower. On GD20, the expression of adrenal IGF1, IGF1R and proliferating cell nuclear antigen (PCNA) were decreased, while the expression of these factors at PW6 were increased in the PCE group. Fetal adrenal gene chip analysis suggested that the mitogen-activated protein kinase/extracellular regulated protein kinase (MAPK/ERK) signal pathway was suppressed in the PCE group. Moreover, in the rat primary adrenal cells, corticosterone (rather than caffeine) was shown to significantly inhibit cell proliferation, IGF1 and PCNA expression, and ERK phosphorylation, which could be reversed by exogenous IGF1. Meanwhile, the effects of exogenous IGF1 were reversed by the ERK pathway inhibitor (PD184161). In conclusion, PCE could induce programming alterations in adrenal cortical cell proliferation before and after birth in male offspring rats. The underlying mechanism is associated with the inhibition of fetal adrenal IGF1-related MAPK/ERK signaling pathway caused by high glucocorticoid levels.

Insulin-like Growth Factor 1 Mediates Adrenal Development Dysfunction in Offspring Rats Induced by Prenatal Food Restriction.

BACKGROUND: Our previous study demonstrated that prenatal food restriction (PFR) could induce the dysfunction of the hypothalamic-pituitary-adrenal axis and glucocorticoid-related glucose and lipid metabolic alterations in adult offspring rats. AIM OF THE STUDY: To investigate the intrauterine programming mechanism of adrenal dysfunction in the PFR offspring rats. METHODS: From gestational days (GDs) 11-20, pregnant Wistar rats were fed a restricted diet (50% of the daily food intake of control rats, 60 g/kg·d). Some were executed at GD20, while the others survived to full-term delivery; all pups were fed a high-fat diet (HFD) after weaning. The serum corticosterone concentration, expression level of adrenal steroidal synthetase, and insulin-like growth factor 1 (IGF1) signaling pathway were tested. RESULTS: We confirmed that the fetal body weight of the PFR group was lower than that of the control group, and the mRNA expression of adrenal steroidogenic acute regulatory protein, cytochrome P450 cholesterol side chain cleavage, 3ß-hydroxysteroid dehydrogenase, and steroid 11ß-hydroxylase (P450c11) were decreased in the PFR fetal rats. The maternal and fetal serum corticosterone levels were significantly increased in the PFR groups. Furthermore, the expression of the adrenal IGF1 signaling pathway (including IGF1, IGF1R, and Akt1) was suppressed. However, after a post-weaning HFD, the body weight gain rates and serum corticosterone levels were elevated, and the expression of adrenal steroid 21-hydroxylase and P450c11, as well as the IGF1 signaling pathway, were significantly increased in the PFR group. CONCLUSIONS: These results showed that a higher level of circulation corticosterone by PFR in utero inhibited adrenal IGF1 signaling and steroidogenesis, whereas post-weaning HFD induced adrenal steroidogenesis by an enhanced IGF1 signaling.

High-fat diet and chronic stress aggravate adrenal function abnormality induced by prenatal caffeine exposure in male offspring rats.

We previously demonstrated thatprenatal caffeine exposure (PCE) suppressed fetal adrenal steroidogenesis and resulted in developmental programming changes in offspring rats. However, whether these changes play a role in adrenal corticosterone synthesis under high-fat diet (HFD) and unpredictable chronic stress (UCS) remains unknown. In present study, rat model was established by PCE (120 mg/kg.d), and male offspring were provided normal diet or HFD after weaning. At postnatal week 21, several rats fed HFD were exposed to UCS for 3 weeks and sacrificed. The results showed that compared with the corresponding control group, the serum corticosterone levels and adrenal steroid synthetase expression of the PCE offspring without UCS were reduced. Moreover, the glucocorticoid (GC)-activation system was inhibited, and insulin-like growth factor 1 (IGF1) signaling pathway expression was increased. With UCS exposure in the PCE offspring, serum corticosterone levels and adrenal steroid synthetase expression were increased, the activity of GC-activation system was enhanced, and adrenal IGF1 signaling pathway expression was decreased. Based on these findings, PCE induced adrenal hypersensitivity in adult male offspring rats, as shown by the reduced corticosterone levels under HFD conditions but significantly enhanced corticosterone levels with UCS, in which GC-IGF1 axis programming alteration may play an important role.

Prenatal caffeine exposure-induced adrenal developmental abnormality in male offspring rats and its possible intrauterine programming mechanisms.

Glucocorticoid (GC) is a major factor for fetal tissue maturation and fate decision after birth. We previously demonstrated that prenatal caffeine exposure (PCE) suppressed fetal adrenal steroidogenesis and resulted in adrenal dysplasia. However, whether these changes play a role until adulthood and its intrauterine programming mechanisms remain unknown. In the present study, a rat model of intrauterine growth retardation (IUGR) was established by PCE, male fetuses and adult offspring were sacrificed at postnatal day (PD) 1, PD7, PD35, PD100 and PD168, respectively. Results showed that the PCE fetal weight decreased and the IUGR rate increased, while the serum corticosterone (CORT) level increased but the insulin-like growth factor 1 (IGF1) level decreased. Fetal adrenal exhibited an enhanced GC-activation system (11ß-hydroxysteroid dehydrogenases/corticoid receptors/CCAAT/enhancer binding proteins), an inhibited IGF1 pathway and steroid synthesis function. After birth, the serum CORT levels in the PCE offspring were increased in the early period followed by falling in the later stage, while the serum IGF1 level change was the opposite and was accompanied by an obvious catch-up growth. Furthermore, the adrenal GC-activation system was inhibited but the IGF1 signaling pathway was enhanced, resulting in a compensatory increase of adrenal steroidogenesis, and the expression of steroidal synthetase was consistent with that of the IGF1 signaling pathway. Based on these findings, we proposed "two-programming mechanisms" for PCE-induced adrenal abnormality: the "first programming" mechanism is a lower function of adrenal steroidogenesis, and prenatal and postnatal adrenal structural and functional abnormalities triggered by the intrauterine GC-IGF1 axis programming-mediated by the GC-activation system that acts as "the second programming" mechanism.

Prenatal ethanol exposure-induced adrenal developmental abnormality of male offspring rats and its possible intrauterine programming mechanisms.

Fetal adrenal developmental status is the major determinant of fetal tissue maturation and offspring growth. We have previously proposed that prenatal ethanol exposure (PEE) suppresses fetal adrenal corticosterone (CORT) synthesis. Here, we focused on PEE-induced adrenal developmental abnormalities of male offspring rats before and after birth, and aimed to explore its intrauterine programming mechanisms. A rat model of intrauterine growth retardation (IUGR) was established by PEE (4g/kg·d). In PEE fetus, increased serum CORT concentration and decreased insulin-like growth factor 1 (IGF1) concentration, with lower bodyweight and structural abnormalities as well as a decreased Ki67 expression (proliferative marker), were observed in the male fetal adrenal cortex. Adrenal glucocorticoid (GC)-metabolic activation system was enhanced while gene expression of IGF1 signaling pathway with steroidogenic acute regulatory protein (StAR), 3ß-hydroxysteroid dehydrogenase (3ß-HSD) was decreased. Furthermore, in the male adult offspring of PEE, serum CORT level was decreased but IGF1 was increased with partial catch-up growth, and Ki67 expression demonstrated no obvious change. Adrenal GC-metabolic activation system was inhibited, while IGF1 signaling pathway and 3ß-HSD was enhanced with the steroidogenic factor 1 (SF1), and StAR was down-regulated in the adult adrenal. Based on these findings, we propose a "two-programming" mechanism for PEE-induced adrenal developmental toxicity: "the first programming" is a lower functional programming of adrenal steroidogenesis, and "the second programming" is GC-metabolic activation system-related GC-IGF1 axis programming.

Intrauterine metabolic programming alteration increased susceptibility to non-alcoholic adult fatty liver disease in prenatal caffeine-exposed rat offspring.

An increase in susceptibility to metabolic syndromes (MetS) in rat offspring that experienced prenatal caffeine exposure (PCE) has been previously demonstrated. The present study aimed to clarify this increased susceptibility and elucidate the mechanism of foetal origin that causes or contributes to adult non-alcoholic fatty liver disease (NAFLD) as a result of PCE. Based on the results from both foetal and adult studies of rats that experienced PCE (120 mg/kgd), the foetal weight and serum triglyceride levels decreased significantly and hepatocellular ultrastructure was altered. Foetal livers exhibited inhibited insulin-like growth factor-1 (IGF-1), enhanced lipogenesis and reduced lipid output. In adult female offspring of PCE+lab chow, lipid synthesis, oxidation and output were enhanced, whereas lipogenesis was inhibited in their male conterparters. Furthermore, in adult offspring of PCE+ high-fat diet, catch-up growth appeared obvious with enhanced hepatic IGF-1, especially in females. Both males and females showed increased lipid synthesis and reduced output, which were accompanied by elevated serum triglyceride. Severe NAFLD appeared with higher Kleiner scores. Gluconeogenesis was continuously enhanced in females. Therefore, increased susceptibility to diet-induced NAFLD in PCE offspring was confirmed, and it appears to be mediated by intrauterine glucose and alterations in lipid metabolic programming. This altered programming enhanced foetal hepatic lipogenesis and reduced lipid output in utero, which continued into the postnatal phase and reappeared in adulthood with the introduction of a high-fat diet, thereby aggravating hepatic lipid accumulation and causing NAFLD.

Mechanisms of anti-inflammatory effect of an active ingredients group from Jinxuan Zhike Xunxi San.

AIM OF THE STUDY: JZXS has been used for inflamed hemorrhoids as a chinese prescription for several decades. The present study was designed to investigate the anti-inflammatory activities, as well as the mechanisms, of an active ingredients group (AIG) obtained from Jinxuan Zhike Xunxi San (JZXS). MATERIALS AND METHODS: The anti-inflammatory activities and mechanisms of AIG were evaluated by xylene-induced ear edema experiments in normal mice and mice without adrenals, leukocyte migration experiments, and carrageenin-induced peritonitis experiments, taking JZXS as the positive control. RESULTS: AIG and JZXS prevented xylene-induced ear edema in normal mice but showed no effects when adrenals were removed. Additionally, AIG and JZXS inhibited leukocyte migration, reduced prostaglandin E2 (PGE2) level in inflammatory exudates and nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin-6 and 8 (IL-6 and IL-8) levels in serum. CONCLUSIONS: AIG and JZXS showed significant anti-inflammatory activities depending on pituitary-adrenal axis, thereby inhibiting leukocyte migration and reducing cytokines and mediators.