Transcriptome analysis reveals that long noncoding RNAs contribute to developmental differences between medium-sized ovarian follicles of Meishan and Duroc sows.

Ovulation rate is an extremely important factor affecting litter size in sows. It differs greatly among pig breeds with different genetic backgrounds. Long non-coding RNAs (lncRNAs) can regulate follicle development, granulosa cell growth, and hormone secretion, which in turn can affect sow litter size. In this study, we identified 3554 lncRNAs and 25,491 mRNAs in M2 follicles of Meishan and Duroc sows. The lncRNA sequence and open reading frame lengths were shorter than mRNAs, and lncRNAs had fewer exons, were less abundant, and more conserved than protein-coding RNAs. Furthermore, 201 lncRNAs were differentially expressed (DE) between breeds, and quantitative trait loci analysis of DE lncRNAs were performed. A total of 127 DE lncRNAs were identified in 119 reproduction trait-related loci. In addition, the potential target genes of lncRNAs in cis or trans configurations were predicted. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed that some potential target genes were involved in follicular development and hormone secretion-related biological processes or pathways, such as progesterone biosynthetic process, estrogen metabolic process, ovarian steroidogenesis, and PI3K-Akt signaling pathway. Furthermore, we also screened 19 differentially expressed lncRNAs in the PI3K-Akt signaling pathway as candidates. This study provides new insights into the roles of lncRNAs in follicular growth and development in pigs.

miR-146a-5p Regulated Cell Proliferation and Apoptosis by Targeting SMAD3 and SMAD4.

BACKGROUND: microRNAs (miRNAs) are a small, endogenous non-coding RNAs that are involved in post-transcriptional gene regulation of many biological processes, including embryo implantation and placental development. In our previous study, miR-146a-5p was found expressed higher in the serum exosomes of pregnant sows than non-pregnant. The research on miR-146a-5p has been mainly related to human diseases, but there are few studies on its effects on the reproduction of sows in early pregnancy. OBJECTIVE: In this article, our motivation is to study the role of miR-146a-5p in the early pregnancy of sows on the cell proliferetion and apoptosis by targeting SMAD3 and SMAD4. METHODS: Bioinformatics software was used to identify the target genes of miR-146a-5p. The wildtype and mutant-type recombinant plasmids of dual-luciferase reporter with 3'-UTR of Smad3 or 3'- UTR of Smad4 were constructed, and co-transfected in porcine kidney cell (PK-15 cell) with miR- 146a-5p mimic, mimic-NC(M-NC), inhibitor and inhibitor-NC(IN-NC), then dual-luciferase activity analysis, qRT-PCR and Western blot were performed to verify the target genes. After the transfection of BeWo choriocarcinoma cell (BeWo cell) with miR-146a-5p mimic, M-NC, inhibitor and IN-NC, the mRNA expression of Caspase-3, BAX and Bcl-2 was measured using qRT-PCR, and the cell proliferation was measured using CCK-8 kit. RESULTS: The luciferase, mRNA and protein expression of Smad3 in PK-15 cells treated by Smad3- 3'-UTR-W co-transfected with miR-146a-5p mimic were significantly lower than that with miR- 146a-5p M-NC, and the results of Smad4 were similar to Smad3, but the protein expression had a trend to lower in mimic group. The expression level of Bcl-2 in the miR-146a-5p mimic group was significantly lower than that in the miR-146a-5p M-NC group, but the expression pattern of Caspase-3 was just opposite. The mimic of miR-146a-5p reduced the proliferation of BeWo cells, however the inhibitor increased. CONCLUSION: Smad3 and Smad4 are the direct target genes of miR-146a-5p. The expression of Smad3 and Smad4 were affected by the mimic and inhibitor of miR-146a-5p. miR-146a-5p affects cell apoptosis and proliferation by regulating their target genes. This study provided new data to understand the regulation mechanism of early pregnancy in sows.

[Clinical observation on the distribution characteristics and rules of pressing sensitive acupoints in bronchial asthma patients].

OBJECTIVE: To compare the distribution characteristics of pressing sensitive acupoints on the body surface between bronchial asthma (BA) patients and healthy subjects, and to analyze the distribution rules of pressing sensitive acupoints in BA patients. METHODS: Seventy BA patients and 70 healthy subjects were selected in this study. The pressing sensitive acupoints were checked with finger pulp and marked on human nerve segment graph. The numbers of pressing sensitive acupoints were counted and the positional relationship between distribution of pressing sensitive acupoints and the position of meridians and nerve segment was observed. RESULTS: (1) The incidence rates of pressing sensitive acupoints in BA patients group and healthy subjects group were 91.4% (64/70) and 15.7% (11/70) respectively, and the BA patients group was higher than the healthy subjects group (P<0.01). (2) The top 3 meridians with pressing sensitive acupoints occuring in BA patients were bladder meridian of foot-taiyang, lung meridian of hand-taiyin and large intestine meridian of hand-yangming, and the most frequent pressing sensitive acupoints were Feishu（BL 13）, Xinshu（BL 15）, Chize（LU 5） and Jueyinshu (BL 14). (3) The pressing sensitive acupoints in BA patients were distributed mainly on C4, C6 and T1-T6 nerve segment. CONCLUSION: Pressing sensitive acupoints have a close correlation with physical condition, and there is a close relation between pressing sensitive acupoints distribution and corresponding meridians and nerve segments in BA patients.

[Distributive regularity of pressing sensitive acupoints in chronic non-atrophic gastritis patients].

OBJECTIVE: To compare the distribution regularity of the pressure-sensitive acupoints in patients with chronic non-atrophic gastritis (CNAG) and healthy subjects, so as to provide suitable acupoint combinations for clinical treatment. METHODS: A total of 120 volunteer subjects including 60 CNAG patients (29 men and 31 women, (40.7±10.3) years at the average age) and 60 healthy subjects (28 men and 32 women, and (40.8±10.2) years at the average age) were enrolled in the present study. The pressure-sensitive acupoints were checked by a fixed operator using his finger pulp along the body trunk and the four limbs and marked on a prepared human dermatome graph. The number of pressing sensitive acupoints were counted, and the relationship between the distribution of the detected sensitive acupoints and the position of meridians and nerve segments was analyzed. RESULTS: The incidence of pressure-sensitive acupoint in CNAG patients and healthy subjects were 86.7% and 15.0%, respectively. In 60 CNAG patients, the most frequently met sensitive acupoints were Xuehai (SP10), Zhongwan (CV13), and Zhongting (CV17) in sequence, mainly covering the Conception Vessel, Spleen Meridian of Foot-Taiyin (SP), and the Stomach Meridian of Foot-Yangming (ST). The sensitive acupoints presented a nerve-segmental distribution within T7-T10 and L3-L5. CONCLUSION: The pressure-sensitive acupoints present a nerve-segmental distribution and have a higher corresponding rate with some meridians related to the stomach, especially under diseased conditions.

[Effects of heat-sensitive moxibustion on HPA axis in rats with irritable bowel syndrome].

OBJECTIVE: To observe the effects of heat-sensitive moxibustion on corticotropin releasing hormone (CRH), adrenocorticotrophic hormone (ACTH) and corticosterone (CORT) in rats with irritable bowel syndrome (IBS), and to explore the possible mechanism of heat-sensitive moxibustion on IBS. METHODS: According to random number table, 56 SD male rats were randomly divided into a blank group (n=8), a model group (n=8), a moxibustion group (n=32), and a mifepristone group (RU-486 group, n=8). The rats in the blank group were treated with normal feeding; the rats in the model group, RU-486 group and moxibustion group were treated with chronic non-predictable stimulation for 21 days to establish IBS model. After model establishment, the rats in the moxibustion group were treated with moxibustion at "Mingmen" (GV 4) for 40 min, once a day for 14 days; the tail temperature was recorded every 5 min; according to the change of tail temperature, the rats were divided into a heat-sensitive moxibustion group and a non-heat-sensitive moxibustion group, and 8 rats were randomly selected in the two groups. The rats in the RU-486 group were treated with gastric administration of RU-486 for 14 days, while the rats in the blank group, model group and moxibustion groups were treated with identical volume of 0.9% NaCl. The rat general condition, body mass, behavioristics, intestinal propulsive rate and visceral sensitivity were observed in each group; ELISA method was used to detect serum CRH, ACTH and CORT; optical microscope was applied to observe the morphological changes of colon. RESULTS: (1) After model establishment, rats were in rest state, fatigued, with withered hair and dim ear; the stool was dry or watery; the body mass were slowly increased; the number of crossed grid and standing frequency were significantly reduced; visceral sensitivity was increased and intestinal propulsion rate was decreased; no obvious inflammatory cell infiltration was observed under microscope. (2) After intervention, compared with the blank group, the body mass and visceral sensitivity in the RU-486 group were not significantly different (both P>0.05), but the intestinal propulsion rate was decreased significantly (P<0.01). Compared with the blank group, the body mass of heat-sensitive moxibustion group and non-heat-sensitive moxibustion group was lower (both P<0.01), but the visceral sensitivity and intestinal propulsion rate were similar (both P>0.05). Compared with the model group, the body mass and visceral sensitivity were improved in the RU-486 group (P<0.05, P<0.01), but the intestinal propulsion rate was similar (P>0.05). The body mass, visceral sensitivity and intestinal propulsion rate of the heat-sensitive moxibustion group and the non-heat-sensitive moxibustion group were superior to those of the model group (P<0.05, P<0.01), and the body mass and intestinal propulsion rate of heat-sensitive moxibustion group were superior to those of non-heat-sensitive moxibustion group (both P<0.05). (3) After intervention, compared with the blank group, the contents of CRH, ACTH and CORT in the model group were significantly increased (P<0.05, P<0.01). Compared with the model group, the contents of CRH, ACTH and CORT of the heat-sensitive moxibustion group were statistically reduced (P<0.05, P<0.01), and the contents of CRH and ACTH in the non-heat-sensitive moxibustion group were statistically reduced (P<0.05, P<0.01); the content of CRH in the RU-486 group was reduced (P<0.05), but the contents of ACTH and CORT were increased (P<0.05, P<0.01). Compared with the non-heat-sensitive moxibustion group, the heat-sensitive moxibustion group was better in the improvement of CRH (P<0.05), but there was no significant difference of ACTH and CORT between the two groups (both P>0.05). CONCLUSION: Heat-sensitive moxibustion could reduce the contents of CRH, ACTH and CORT through the HPA axis, and improve the function of gastrointestinal motility to treat IBS.