Molecular cloning, sequence, and expression patterns of DNA damage induced transcript 3 (DDIT3) gene in female yaks (Bos grunniens).

Endoplasmic reticulum stress (ERS) plays an important role in regulating the reproductive process of female mammals, mainly involved in follicular atresia and corpus luteum regression. DNA damage induced transcript 3 (DDIT3) is a marker gene of ERS. The objectives of the present study were to clone and analyze the sequence and tissue expression characteristics of DDIT3 gene in female yaks. By reverse transcriptase-polymerase chain reaction (RT-PCR) strategy, we obtained full-length 507-bp DDIT3-cDNA, encoding for 168-aa protein. Yak DDIT3 exhibited highest and least identity with that of bison and horse, respectively. Real-time PCR analyses revealed that the expression level of DDIT3 gene in ovary was higher than that in heart, liver, kidney, spleen, lung, uterus and oviduct (p < 0.05). DDIT3 expression level in ovary and uterus during pregnancy was higher than that in follicular phase, luteal phase and fetus stage. DDIT3 was highly expressed in metaphase II oocytes and granulosa cells than that in germinal vesicle and metaphase I oocytes (p < 0.05), respectively. This is the first molecular characterization and expression patterns of DDIT3 gene in female yaks. These results indicated that the DDIT3 gene possibly plays an important role in regulating ovary function and pregnancy maintenance in yaks.

Comparison of the sequences and expression levels of genes related to follicular development and atresia between prolific and nonprolific goat breeds.

This study investigated the variations of the nucleotide sequences and ovarian expression levels of genes related to follicular development and atresia in prolific Jintang black goats and nonprolific Tibetan goats. Eight genes, FSHB, LHB, FSHR, LHCGR, ESR2, B4GANT2, BCL2 and BAX, were examined using reverse transcription-polymerase chain reaction and quantitative real-time PCR. The results showed that the nucleotide and deduced amino acid sequences of the LHB and BAX genes were not different, but there was one base change in the FSHR genes between the two breeds. There was one base change in the FSHB gene, which resulted in one amino acid substitution; there were nine base changes in the LHCGR gene, which resulted in five amino acid substitutions; and there were six base changes in the B4GANT2 gene, which resulted in four amino acid substitutions. The expression levels of the FSHR, LHCGR, ESR2, B4GANT2, BCL2 and BAX genes in the ovaries were not different between the two breeds. The plasma concentrations of FSH were not different, but the plasma concentrations of LH, P4 and E2 were lower in prolific Jintang black goats than in nonprolific Tibetan goats (P ˂ 0.05) at 40 hr after removal of the Controlled Internal Drug Release Devices. These results provide some foundations elucidating the endocrine and molecular mechanisms controlling ovulation rate in goats, but these need to be further verified.

Identification and comparative analysis of the ovarian microRNAs of prolific and non-prolific goats during the follicular phase using high-throughput sequencing.

The kidding rate is one of the most important economic traits for goat production, but the genetic mechanism that is associated with ovulation rate is poorly understood. Recently, increasing evidence has suggested that microRNAs (miRNAs) influence ovarian biological processes. The present study provides the first comparison of the ovarian miRNAs of prolific Jintang black goats (JTGs) and non-prolific Tibetan goats (TBGs) during the follicular phase using RNA-Seq technology. We generated 11.19 million (M) and 11.34 M clean reads from the TBG and JTG libraries, respectively, from which a total of 389 known miRNAs were identified and 142 novel miRNAs were predicted. A total of 191 miRNAs were differentially expressed between the two breeds. Among the 10 most abundant miRNAs, miR-21-5p was defined as differentially expressed miRNA with a higher level in the JTG library than in the TBG library, but the other miRNAs were not different between the breeds. The predicted miRNA-targeted genes were further analyzed by Gene Ontology and KEGG pathway analyses. The results revealed that miR-21, miR-99a, miRNA-143, let-7f, miR-493 and miR-200b may affect follicular development. These findings will increase the current understanding of the role of ovarian miRNAs in the regulation of ovulation rate in goats.

Comparative messenger RNA expression of FSHß, LHß, FSHR, LHR, and ERß in high and low prolific goat breeds.

Follicle-stimulating hormone (FSH) and luteinizing hormone (LH) have a central role in follicle growth and maturation, but no clear differences between breeds with different ovulation rates have been found. Therefore, this study investigated mRNA expression of FSHß, LHß, FSH receptor (FSHR), LH receptor (LHR), and estrogen receptor-ß (ERß) genes in prolific Lezhi black (LB) goats and nonprolific Tibetan (TB) goats by real-time PCR. Follicles and pituitaries were recovered from goats at 12-24 h after onset of estrus. Real-time PCR analysis revealed that the expression levels of FSHß and LHß mRNA were significantly higher (p < 0.01) in pituitary of LB than in TB does, but the expression levels of FSHR and LHR mRNA in follicle of TB were greater (p < 0.05). Expression level of follicular ER ß was not different between the two breeds. Data provide evidence that the greater ovulation rate in the LB goat as compared to the TB breed is associated with a greater gonadotropin expression during follicular phase.

[Expansive pedicle screws fixation combined with Wright artificial bone implantation for treatment of thoracolumbar burst fracture of the elderly patients].

OBJECTIVE: To observe the effects of expansive pedicle screws fixation combined Wright artificial bone implantation in treating thoracolumbar burst fracture of the elderly. METHODS: From March 2007 to July 2009, 12 patients with thoracolumbar burst fractures were treated with surgery in the study. There were 7 males and 5 females, with an average age of 62.3 years (from 56 to 71 years). Nerve functions of all patients were ASIA classified grade E. According to TLICS score system, 6 points were in 5 cases and 7 points were in 7 cases. Expansive pedicle screw system was used and Wright artificial bone was injected into the injuried vertebrae. The anterior border height of vertebral body and Cobb angle were observed at the preoperation, postoperation and final follow-up. RESULTS: All patients were followed up from 10 to 15 months with an average of 12 months. Wounds healed well, pain in the chest-back abated, no the expansive pedicle screws loosened or broke down. The anterior border height of vertebral body increased from (32.3 +/- 9.1)% preoperatively to (95.3 +/- 3.2)% postoperatively; and the Cobb angle decreased from (31.6 +/- 6.8) degrees preoperatively to (4.5 +/- 3.2) degrees postoperatively. There was significant difference between two groups (P<0.01). After 3 months, the anterior border height of vertebral body was (94.7 +/- 3.3)% and the Cobb angle was (4.6 +/- 3.4) degrees; at the final follow-up, the anterior border height of vertebral body was (93.2 +/- 3.6)% and the Cobb angle was (5.3 +/- 3.7) degrees. There was no significant difference between the two data (P>0.05). CONCLUSION: The combination of expansive pedicle screws fixation and Wright artificial bone injection is a good treatment for thoracolumbar burst fracture, with advantages such as easy operation and good effect, which mainly applied to patients with no obvious nerve injuries.

[Clinical observation of anterior cruciate ligament reconstruction with tibial-remnant preserving technique using hamstring autograft].

OBJECTIVE: To observe the outcome of ACL reconstruction with tibial-remnant preserving technique using hamstring autograft. METHODS: From January 2007 to June 2009,18 patients, including 11 male and 7 female with an average age of 28.2 years (from 18 to 42 years), received ACL reconstruction in Kunshan traditional Chinese medicine hospital, Jiangsu province. All cases were confirmed that the remnant of torn ACL were more than 1/3 by arthroscopic observation. The gracillis and semitendinosus tendons harvested from the patient were folded and weaved as a graft, which was pulled into the femoral and tibia tunnel through the sheath formed by the remnant preserved carefully, then fixed by Rigidfix at femoral side and Intrafix at tibial side respectively. The Lysholm scores were recorded at the last follow-up to evaluate the postoperative knee function. The knee stability, range of motion (ROM), and proprioception were evaluated by physical examination. RESULTS: The average duration of follow-up of all 18 patients was 17.3 months (ranged from 15 to 24 months), no synovitis, graft tear, and ROM limitation were found. The average postoperative Lysholm score was (91.56 +/- 4.60) improved from preoperative (49.43 +/- 2.85) (P < 0.05). There were no differences between bilateral knees on the knee flexion (P > 0.05). CONCLUSION: To preserve the tibial remnant of torn ACL during ACL reconstruction had good outcomes, and contributed to recovery of knee proprioceptive function.

Induction of estrus in suckled female yaks (Bos grunniens) and synchronization of ovulation in the non-sucklers for timed artificial insemination using progesterone treatments and Co-Synch regimens.

The objectives of the present study were to evaluate the induction of estrus and fertility in yak cows treated with Co-Synch regimens or progesterone (P(4)). In Experiment 1, postpartum suckled yaks were assigned to three treatments: (1) A (n=28), insertion of an intravaginal device containing P(4) (CIDR) on Day 0, PGF(2alpha) (i.m.) on Day 6 and PMSG (i.m.) at the time of CIDR removal on Day 7 (P(4)-PGF(2alpha)-PMSG); (2) B (n=21), PGF(2alpha) (i.m.) on Day 6 and PMSG on Day 7; (3) C (n=26), control group. Seven yak bulls were grazed with the cows for natural breeding. Rate of estrus within 96h of the end of treatment was greater (P<0.05) in A (100.0%) than in B (28.6%) or C (0.0%). First service conception rate (CR) determined by serum P(4) on Day 21 after breeding was greater (P<0.05) in A (78.6%) than in B (22.2%). Also, pregnancy rate (PR) during the breeding season was greater (P<0.05) in A (82.1%) than in B (19.0%) and C (7.7%). In Experiment 2, non-suckled yaks that calved in previous years but not in the current year were assigned to three treatments: (1) A (n=31), GnRH (i.m.) on Day 0, followed by PGF(2alpha) on Day 7 and timed artificial insemination (TAI) concurrently with GnRH treatment on Day 9 (Co-Synch regimen); (2) B (n=50), a CIDR device for 7 days plus PGF(2alpha) and PMSG at the time of CIDR withdrawal on Day 7 and TAI on Day 9 (P(4)-PGF(2alpha)-PMSG); (3) C (n=50), yak cows were artificially inseminated at spontaneous estrus. Frozen semen of Holstein and Jersey were used for insemination in Experiment 2. The CR assessed by rectal palpation 35 days after TAI was not different in A (22.6%), B (30.0%) and C (33.3%), but PR was greater in A and B than in C, when based on those cows presented for estrous synchronization programs. It is concluded that P(4)-PGF(2alpha)-PMSG protocol could efficiently induce estrus and result in an acceptable pregnancy rate in postpartum suckled yak cows. This technique and Co-Synch regimen can be applied successfully for TAI of non-suckled yak cows.