The Ziwuling black goat is an indigenously in China, their offspring are frequently affected by congenital cryptorchidism. The extracellular matrix (ECM) contains cytokines and growth factors that regulate the development of the testis, and component changes often result in pathological changes. Cryptorchidism is closely related to structural changes in ECM. In this study, the histochemical staining, immunohistochemical, immunofluorescence and Western blot combined with semi-quantitative analysis was used to describe the distribution of the important ECM components Collagen type IV (Col IV), laminin (LN)and heparan sulfate proteoglycans (HSPG) in the normal and cryptorchid testes of Ziwuling black goats. Results showed that: The histochemical staining showed that the dysplasia of seminiferous tubules and decreased number of Sertoli cells in cryptorchidism, as well as sparse collagen fiber. Meanwhile, the distribution of reticular fibers is relatively rich. Furthermore, the PAS and AB staining in the interstitial vessels and lamina propria of seminiferous tubules is weak. The immunohistochemical and immunofluorescence revealed that Col IV, LN was strongly expressed in Leydig, Sertoli cells of normal testes and moderately positive in the spermatogonia and spermatids, but HSPG was not expressed in the spermatogonia. However, cryptorchidism, the expression of Col IV, LN and HPSG in Leydig, Sertoli cells significantly decreased, as well as the expression of Col IV and LN in capillary endothelial cells, but HSPG was moderately expressed in spermatogonia. Based on these data, the underdevelopment of spermatogenic epithelium, decreased synthesis function of collagen fibers and Leydig cells develop usually in the cryptorchidism were shown to be closely related to the abnormal metabolism of Col IV and LN. The positive expressed of HSPG in the spermatogonia of cryptorchid testes is related to the compensatory development of spermatogonia.

Escherichia coli (E. coli) is one of the major pathogenic bacteria in bovine mastitis, which usually triggers systemic symptoms by releasing lipopolysaccharide (LPS). waaF is the core in LPS pathogenicity. In this study, a new waaF vaccine candidate was identified, constructed with the pcDNA3.1 (+)HisB-waaF plasmid to create to a DNA vaccine (pcwaaF), and transfected into MCF-7 cells to produce recombinant waaF subunit vaccine (rwaaF). After that, the safety of the two vaccine candidates was evaluated in mouse model. Immunogenicity and mortality of challenged mice were compared in 20 and 40 μg per dose, respectively. The results showed that rwaaF and pcwaaF were successfully constructed and the complete blood count and serum biochemical indicated that both of the vaccine candidates were safe (p > 0.05). In addition, histopathological staining showed no obvious pathological changes. The immune response induced by rwaaF was significantly higher than that of pcwaaF (p < 0.01), indicated by levels of serum concentration of IgG IL-2, IL-4, and IFN-γ, and feces concentration of sIgA. Survival rates of mice in rwaaF groups (both 80%) were also higher than in the pcwaaF groups (40 and 50%, respectively). Comparing the safety, immunogenicity, and E. coli challenge of two vaccine candidates, rwaaF had the better effect and 20 μg rwaaF was more economical. In conclusion, this study demonstrates the utility of a new E. coli vaccine and provides a rationale for further investigation of bovine mastitis therapy and management.

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This study aimed to investigate the distribution and expression of matrix metalloproteinase‐2 (MMP‐2) and tissue inhibitor of matrix metalloproteinase‐2 (TIMP‐2) in yak testes. The testes of healthy yaks at different ages: newborn [3 days], young [1 year], adult [4 years], and old [9 years] were collected for microscopic analyses using hematoxylin and eosin staining, immunohistochemistry and immunofluorescence, as well as western blot to compare the expression of MMP‐2 and TIMP‐2. Furthermore, the levels of MMP‐2mRNA and TIMP‐2mRNA was detected by real‐time quantitative polymerase chain reaction (qPCR). The results of immunohistochemistry and immunofluorescence demonstrated that MMP‐2 and TIMP‐2 were mainly located in gonocytes of newborn, Sertoli cells of young, spermatozoa of adult and Leydig cells of old. The protein levels of MMP‐2 and TIMP‐2 exhibited a downward from newborn to adult, but increased again in old yaks. The analysis of qPCR showed that MMP‐2 was higher in young compared with newborn or adult(**p < .01), but a lower expression was detected in adult compared with old yak testicular tissues (*p < .05). Compared with adults, TIMP‐2 was significantly higher in newborn and young yaks (**p < .01), and slightly higher in old yaks (*p < .05). Hence, The location of MMP‐2 and TIMP‐2 in gonocytes were associated with the development of newborn yak testes. The expression of MMP‐2 and TIMP‐2 in Sertoli cells at young and adult yaks suggested that they provided a clue for the regulation of spermatogenesis. The positive labeling of MMP‐2 and TIMP‐2 in Leydig cells in old yaks suggested that both may be involved in the interstitial metabolism of the testes during this period. This study revealed the possible role of MMP‐2 and TIMP‐2 in testicular functionality of yaks at different ages.