[PubMed] [Google Scholar] 3. FSHR-57aa was significantly decreased while the serum hormone levels of testosterone and estradiol were not disturbed in the control or the FSHR-57aa groups. By evaluating pathological changes in testicular 4-O-Caffeoylquinic acid histology, we found that the blood-testis barrier remained intact, in spite of some small damage to Sertoli cells. In conclusion, our study demonstrates that this rhFSHR-57aa protein might be a feasible male contraceptive which could impact sperm production without disturbing hormone levels. strain targeted the FSHR N-terminal 57aa sequence and overlapping 18C74 residues was designed and prepared, and its immune effects in the rhesus monkey were observed. MATERIALS AND METHODS Antigenic epitope prediction and plasmid construction Antigenic epitopes of the human FSHR (GenBank No. “type”:”entrez-protein”,”attrs”:”text”:”AAI25271″,”term_id”:”115940701″,”term_text”:”AAI25271″AAI25271) were predicted by DNASTAR software (DNASTAR Inc., Madison, USA) from your characteristics of its antigenic index analysis. Sequence alignment and homology comparisons were performed 4-O-Caffeoylquinic acid around the homology between the FSHR and LHR proteins (LHR, GenBank No. “type”:”entrez-protein”,”attrs”:”text”:”NP_000224″,”term_id”:”106067657″,”term_text”:”NP_000224″NP_000224) to avoid the design of a protein that may cross-talk with LHR. Total RNA was extracted from a human testis (three adult testes; 32, 33 and 35-years-old, stored at ? 70C for a month) by using TRIzol reagent and reverse transcribed to cDNA according to the manufacturer’s protocol. The FSHR-57aa fragment (GeneBank No. “type”:”entrez-nucleotide”,”attrs”:”text”:”NM_181446″,”term_id”:”1677531548″,”term_text”:”NM_181446″NM_181446) (nucleotides 162C332) was amplified by polymerase chain reaction with the following primers: 5-GTTC-CCATGG (yeast extract, 1% (SDS-polyacrylamide gel electrophoresis (SDS-PAGE; 4C, 100 V and 15 h). The gel slices containing FSHR-57aa proteins (7 kDa) were cut out and put into a dialysis sac with 3.5 ml electrophoresis buffer. The sac underwent horizontal electrophoresis 4-O-Caffeoylquinic acid at 150 V for 2 h at 4C and then reversed-pole electrophoresis for 5 min. The buffer in the dialysis sac with target FSHR-57aa proteins was collected. The dialysis-electrophoresis process was repeated and about 7 ml protein sample was harvested. 0.05. Data were analyzed by independent-samples and -0.01 (a). Multiple comparisons made by the Games-Howell test indicated that after 6 weeks antibody titres were different from those in the untreated state (0 week), * 0.05, ** 0.01. The antibody titre in seminal plasma (b). The data are expressed as the mean s.d. of three impartial experiments. The antibody titre in seminal plasma, when the dilution was 1:100, showed a slight but nonsignificant increase on day 140 (20th week) after first immunization (Physique 5b). Fertility evaluation The pregnancy result is shown in Table 2. The pregnancy rates of females in the control and FSHR-57 groups were 94.4% and 61.1%, respectively, with the chi-squared test = 5.8, and Fisher’s exact value was 0.041. The sperm parameter values are shown in Physique 6 as total concentration (106 ml?1), motion (%) and linearity (%) of the FSHR-57 and control groups. The total sperm concentration of the control and FSHR-57 Rabbit polyclonal to KATNB1 groups was 578.8 281.86 (106 ml?1) and 419.5 204.3 (106 ml?1), respectively (Physique 6a). Sperm motion of the control and FSHR-57 groups was 70.2% 10.9% and 60.9% 7.9%, respectively (Determine 6b), and the linearity of the control and FSHR-57 group was 45.4 14.8 and 27.8 8.3, respectively 4-O-Caffeoylquinic acid (Determine 6c). All of results represent the mean standard error of the mean. The value of sperm parameter values were all 0.05. Although serum levels of testosterone and estradiol ranged widely within and between monkeys at different times, no significant statistical differences were observed in them between the FSHR-57aa and control group (Physique 7). In the FSHR-57aa protein immunized group, we did not observe obvious degeneration of testicular tissue. However, some slight changes were found in the FSHR-57aa immunized group, such as apparent slight damage to Sertoli cells and a possible decreased quantity of round spermatids and spermatozoa in the seminiferous tubules (Physique 8). Morphological quantification indicated that this diameters of the seminiferous tubules was significantly different between the two groups, while there were no significant changes in the thickness of seminiferous epithelium (Table 3). Table 2 Fertility evaluation Open in a separate window Open in a separate window Physique 6 The sperm parameters were compared in two groups..