Experimental Biology and Medicine

Chronic alcohol intake causes hepatic steatosis and changes the body composition and glucose metabolism. We examined whether water extracts of mulberry (WMB) and white flower dandelion (Taraxacum coreanum Nakai, WTC) can prevent and/or delay the symptoms of chronic ethanol-induced hepatic steatosis in male Sprague Dawley rats, and explored the mechanisms. Ethanol degradation was examined by orally administering 3 g ethanol/kg bw after giving them 0.3 g/kg bw WMB or WTC. All rats were continuously provided about 7 g ethanol/kg bw/day for four weeks and were given either of 0.1% dextrin (control), WMB, WTC, or water extracts of Hovenia dulcis Thunb fruit (positive-control) in high-fat diets. Area under the curve of serum ethanol levels was lowered in descending order of control, WTC and positive-control, and WMB in acute ethanol challenge. WMB and WTC prevented alcohol intake-related decrease in bone mineral density and lean body mass compared to the control. After glucose challenge, serum glucose levels increased more in the control group than other groups in the first part and the rate of decrease after 40 min was similar among all groups. These changes were associated with decreasing serum insulin levels. WMB had the greatest efficacy for decreasing triglyceride and increasing glycogen deposits. WMB and WTC prevented the disruption of the hepatic cells and nuclei while reducing malondialdehyde contents in rats fed alcohol, but the prevention was not as much as the normal-control. The ratio of Firmicutes to Bacteroidetes in the gut was much higher in the control than the normal-control, but WTC and WMB decreased the ratio compared to the control. WMB and WTC separated the gut microbiota community from the control. In conclusion, WMB and WTC protected against alcoholic liver steatosis by accelerating ethanol degradation and also improved body composition and glucose metabolism while alleviating the dysbiosis of gut microbiome by chronic alcohol intake.

Dronedarone improves microvascular flow during atrial fibrillation and reduces the infarct size in acute models of myocardial infarction. However, dronedarone might be harmful in patients with recent decompensated heart failure and increases mortality in patients with permanent atrial fibrillation. A pathophysiological explanation for these discrepant data is lacking. This study investigated the effects of dronedarone on gene and protein expression in the infarcted area and border zone in pigs subjected to anterior ischemia/reperfusion myocardial infarction. The ischemia/reperfusion myocardial infarction was induced in 16 pigs. Eight pigs were treated with dronedarone for 28 days after myocardial infarction, the remaining pigs served as control. Microarray-based transcriptome profiling and 2D-DIGE-based proteome analysis were used to assess the effects of dronedarone on left ventricular gene expression in healthy (LV), infarcted (MI), and border zone tissue. Selected targets were validated by RT-qPCR or immunoblot analyses, with special emphasize given to the transcriptome/proteome overlap. Combined “omics” analysis was performed to identify most significant disease and function charts affected by dronedarone and to establish an integrated network. The levels of 879 (BZ) or 7 (MI) transcripts and 51 (LV) or 15 (BZ) proteins were significantly altered by dronedarone, pointing to a substantial efficacy of dronedarone in the border zone. Transcriptome and proteome data indicate that dronedarone influences post-infarction remodeling processes and identify matricellular proteins as major targets of dronedarone in this setting. This finding is fully supported by the disease and function charts as well as by the integrated network established by combined “omics”. Dronedarone therapy alters myocardial gene expression after acute myocardial infarction with pronounced effects in the border zone. Dronedarone promotes infarct healing via regulation of periostin and might contribute to the limitation of its expansion as well as cardiac rupture. Thus, there are no experimental hints that dronedarone per se has direct harmful effects after MI in ventricular tissue.

The intestinal epithelium is continuously regenerated through proliferation and differentiation of stem cells located in the intestinal crypts. Obesity affects this process and results in greater stem cell proliferation and altered tissue growth and function. Obesity-induced high levels of insulin and insulin-like growth factor-1 in the stem cell niche are found to impact proliferation in rodents indicating that insulin and insulin-like growth factor-1 receptors may play a role in modulating intestinal epithelial stem cell proliferation. To determine whether insulin or insulin-like growth factor-1 can induce proliferation in human intestinal epithelial stem cells, and if two downstream insulin and insulin-like growth factor-1 receptor signaling pathways, PI3K/Akt and ERK, are involved, we used primary small intestinal epithelial crypts isolated from obese humans and investigated (1) the effect of insulin or insulin-like growth factor-1 on crypt proliferation, and (2) the effect of insulin and insulin-like growth factor-1 signaling inhibitors on insulin or insulin-like growth factor-1-induced proliferation. We found that insulin and insulin-like growth factor-1 enhanced the proliferation of crypt cells, including intestinal epithelial stem cells. Inhibition of the PI3K/Akt pathway attenuated insulin and insulin-like growth factor-1-induced proliferation, but inhibition of the ERK pathway had no effect. These results suggest that the classical metabolic PI3K pathway and not the canonical proliferation ERK pathway is involved in the insulin/insulin-like growth factor-1-induced increase in crypt proliferation in obese humans, which may contribute to abnormal tissue renewal and function.

The chromatoid body is a cytoplasmic male germ cell structure that plays a role in the regulation of mRNA transcription during spermatogenesis. A proteomic analysis of this structure has identified the presence of its classic molecular markers (MVH and MIWI), as well as a significant number of transient proteins. Circadian locomotor output cycles protein kaput (CLOCK) and brain and muscle ARNT-like 1 (BMAL1), which are molecular components of the circadian clock, are likely located in the chromatoid body in a transient fashion. This study sought to determine whether aging produces morphological changes in the chromatoid bodies of round spermatids similar to those previously observed in BMAL1 knockout mice. A sample of 30 male mice was divided into three groups: juvenile mice (45 days old), adult mice (120 days old), and old mice (+180 days old). Aging was confirmed by viability and sperm count analyses and testosterone dosage. Squash slides prepared with fragments of seminiferous tubules were immunostained for MVH, MIWI, BMAL1, and CLOCK detection. In juvenile and adult specimens, single round chromatoid bodies were observed using MVH/BMAL1 and MIWI/CLOCK immunostaining. In old specimens, many chromatoid bodies displayed changes in number and morphology, as well as an increase in the interactions between MVH and BMAL1; MIWI and CLOCK. Changes in chromatoid body morphology increased interactions between the proteins analyzed herein, and decreased amounts of these proteins in seminiferous tubules of older mice may indicate that aging influences the assembly and physiology of chromatoid bodies, which may, in turn, affect fertility.

Hyperhomocysteinemia (HHcy) is associated with suppressed lipolytic response in adipocytes/adipose tissue, however, the underlying mechanism remains to be extensively studied. Nuclear factor erythroid 2-related factor 2 (Nrf2), a master transcriptional factor regulating antioxidant generation, has been recently reported to mediate lipid metabolism. Employing both fully differentiated 3T3-L1 adipocytes and male C57BL/6 mice, in the present study, we investigated the potential involvement of Nrf2 activation in HHcy-mediated lipolytic suppression. Our results showed that homocysteine (Hcy) treatment resulted in suppressed lipolysis, evidenced by increased intracellular triglyceride (TG) accumulation, decreased glycerol and free fatty acid (FFA) in fully differentiated 3T3-L1 adipocytes. Interestingly, Hcy exposure was associated with Nrf2 activation in adipocytes. Further studies showed that Nrf2 knockdown via siRNA transfection ameliorated Hcy-induced glycerol release in adipocytes. On the contrary, Nrf2 activators, epigallocatechin gallate (EGCG) and tert-butylhydroquinone (t-BHQ), increased intracellular TG content and decreased glycerol release in adipocytes. Importantly, our in vitro observations were corroborated by our in vivo findings, in which Hcy feeding (0.1% wt/vol) for four weeks induced Nrf2 expression in adipose tissue and lowered circulating FFA and glycerol levels in mice. Furthermore, EGCG injection (5 mg/kg/d) decreased circulating glycerol levels in comparison to the control group in mice. In conclusion, these results indicated that Nrf2 activation in response to HHcy plays an important role in mediating Hcy-suppressed lipolysis in adipocytes.

Severe lung damage is a major cause of death in blast victims, but the mechanisms of pulmonary blast injury are not well understood. Therefore, it is important to study the injury mechanism of pulmonary blast injury. A model of lung injury induced by blast exposure was established by using a simulation blast device. The effectiveness and reproducibility of the device were investigated. Eighty mice were randomly divided into eight groups: control group and 3 h, 6 h, 12 h, 24 h, 48 h, 7 days and 14 days post blast. The explosive device induced an explosion injury model of a single lung injury in mice. The success rate of the model was as high as 90%, and the degree of lung injury was basically the same under the same pressure. Under the same conditions, the thickness of the aluminum film can be from 0.8 mm to 1.6 mm, and the peak pressure could be from 95.85 ± 15.61 PSI to 423.32 ± 11.64 PSI. There is no statistical difference in intragroup comparison. A follow-up lung injury experiment using an aluminum film thickness of 1.4 mm showed a pressure of 337.46 ± 18.30 PSI induced a mortality rate of approximately 23.2%. Compared with the control group (372 ± 23 times/min, 85.9 ± 9.4 mmHg, 4.34 ± 0.09), blast exposed mice had decreased heart rate (283 ± 21 times/min) and blood pressure (73.6 ± 3.6 mmHg), and increased lung wet/dry weight ratio(2.67 ± 0.11), marked edematous lung tissue, ruptured blood vessels, infiltrating inflammatory cells, increased NF-κB (4.13 ± 0.01), TNF-α (4.13 ± 0.01), IL-1β (2.43 ± 0.01) and IL-6 (4.65 ± 0.01) mRNA and protein, decreased IL-10(0.18 ± 0.02) mRNA and protein (P < 0.05). The formation of ROS and the expression of MDA5 (4.46 ± 0.01) and IREα (3.43 ± 0.00) mRNA and protein were increased and the expression of SOD-1 (0.28 ± 0.02) mRNA and protein was decreased (P < 0.05). Increased expression of Bax (3.54 ± 0.00) and caspase 3 (4.18 ± 0.01) mRNA and protein inhibited the expression of Bcl-2 (0.39 ± 0.02) mRNA and protein. The changes of pulmonary edema, inflammatory cell infiltration, and cell damage factor expression increased gradually with time, and reached the peak at 12–24 h after the outbreak, and returned to normal at 7–14 days. Detonation injury can lead to edema of lung tissue, pulmonary hemorrhage, rupture of pulmonary vessels, induction of early inflammatory responses accompanied by increased oxidative stress in lung tissue cells and increased apoptosis in mice experiencing blast injury. The above results are consistent with those reported in other literatures. It is showed that the mouse lung blast injury model is successfully modeled, and the device can be used for the study of pulmonary blast injury.

Previously, we reported that orally administered Emu Oil (EO) increases mucosal thickness in the small intestine and colon in rodent models of chemotherapy-induced mucositis and colitis. However, it remains unclear whether mucosal thickening (crypt and villus lengthening) represents a process of normal or aberrant growth. We sought to determine if villus height (VH) and crypt depth (CD) measurements returned to normal in EO-treated rats following withdrawal of EO therapy. Dark agouti rats (n = 8/group) were gavaged daily for 10 days with water, olive oil (OO), or EO (0.5 mL or 1 mL). Groups of rats were euthanized on days 10 and 17. Intestinal weights, lengths, VH, and CD were quantified. P < 0.05 was considered significant. On day 10, jejuno–ileum weight was increased by OO (26%) and EO (0.5 mL: 15%; 1 mL: 29%) compared to water controls (P < 0.01), which was normalized by day 17. On days 10 and 17, jejuno-ileum length was greater in OO- (12%) and EO-treated rats (0.5 mL: 8%; 1 mL: 12%; P < 0.05), relative to water controls. On day 10, OO and EO increased ileal VH (OO: 32%; 0.5 EO: 22%; EO: 35%; P < 0.01) and CD (OO: 17%; 0.5 EO: 13%; EO: 22%) compared to water controls. Importantly, however, after withdrawal of all oils, VH and CD measurements returned to normal control values. Moreover, the VH:CD ratio (potential indicator of dysplasia) remained unchanged in all experimental groups on days 10 and 17. The restoration of normal intestinal architecture following cessation of Emu Oil therapy supports its safety for application in intestinal disorders.

Endometriosis, characterized by the presence of endometrial tissue at ectopic sites, is a leading cause of pelvic pain and subfertility in women. The stromal compartment of the endometrium is considered to play a pivotal role in the establishment and persistence of endometriotic lesions, thus impaired decidualization of these cells may result in enhanced invasion capacity at ectopic sites. Consequently, stimulation of decidualization may alleviate this disease. To analyze the effect of systemically applied compounds on decidualization of ectopic endometrial tissue, endometriosis was induced by suturing human eutopic endometrium to the peritoneum of 22 NOD/SCID mice. Each mouse received four tissue fragments from the same patient. Mice were randomly allocated either to one control and three experimental groups (n = 4/group) which were treated with progesterone alone or in combination with forskolin or human chorionic gonadotropin for seven days or to one control and one experimental group (n = 3/group) which was treated with progesterone and human chorionic gonadotropin for 10 days followed by 7 days without treatment. At the end of the experiments, lesions were measured and analyzed for markers of decidualization (FOXO-1, prolactin) and proliferation (Ki-67). Decidualization was induced in the ectopic lesions by systemic treatment in vivo. This induction was significantly stronger after treatment with progesterone in combination with human chorionic gonadotropin than with forskolin or with progesterone alone. Only the combination with human chorionic gonadotropin led to induction of FOXO1 protein expression and a significant physiologic transformation of the ectopic endometrial stromal cells after seven days of treatment. After termination of human chorionic gonadotropin treatment, the decidualization process continued, leading to a significant inhibition of proliferation. Thus, decidualization of human ectopic endometrial tissue can be induced in a humanized endometriosis mouse model in vivo. This model may help to decipher the signal pathways involved in this decidualization process and to develop novel therapeutical approaches to alleviate this painful disease.

Liu J, Fei L, Huang G-Q, Shang X-K, Liu M, Pei Z-J, Zhang Y-X. Right ventricle performances with echocardiography and ^99mTc myocardial perfusion imaging in pulmonary arterial hypertension patients. Exp Biol Med 2018;243(9):754-761.

In this article, the first author’s affiliation was incorrect in the OnlineFirst and print versions. It should have appeared as ‘Department of Nuclear Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China’. This has been corrected in the online article.