| The poultry industry in China has shown tremendous advancement in last three decades, mainly due to the induction of technological interferences. This has led China as the largest producer of eggs and meat in the world. In addition, industry is also fostering the nutritional requirements of 1.35 billion peoples of the country with good quality proteins. Mainly, poultry industry of China is dependent on chicken farming which is dependent on the meat type and egg type breeds from the local and imported stocks. Chicken eggs being rich in biological value and a source of vitamins, minerals and proteins are cheap and easy to consume in variety of forms. However, due to multifold rising economy of the country, the buying ability of the consumers is increasing which is exerting heavy burden on the poultry industry. It is also evident from FAO next ten years projections for per capita chicken meat consumption to increase from 11.2 to 13.8 per Kg per annum. Obviously; it will be hard to cope with the supply and demand for poultry products with the low producing local breeds, which are mainly preferred due to the consumer’s traditional preferences. This can only be achieved by identifying the biological factors or characters that hamper the productive efficiency of the local breeds in comparison to the high producing breeds and then, compensating the deficient characters in the present breeds by breeding programs. Since the major aspect of the production performance are related with the reproductive efficiency which depends on the follicular development, therefore, in-depth research work is required to understand the mechanisms that regulate their development and to manipulate them for the betterment of the breeds.Keeping in view the above scenario, we have compared traditional hens with the commercial hens and cultured follicles, and investigated the factor that can affect the follicular growth in hens.Our main results are described below.1. Comparison of high and low producing breedsIn the first experiment we compared egg forming organs of the high and low producing hens, in order to understand their morphological differences between two different breeds of hens.For this purpose a case study was undertaken on Chinese breed Zhenning hen, which is the cross of the multiple strains of the original Ninghaimeilinji breed, commonly found in the Zhejiang province P.R. China, has been selected as a traditional breed to compare with the Hyline hens. The hens were selected to compare organ body weight ratios in liver, ovary and oviduct. Moreover histomorphology of the liver, ovary and oviduct along with its regions between two breeds was also compared.In addition,egg quality of the two breeds was also compared. The results showed that the liver, ovary and oviduct body weight ratios were significantly higher in the Hyline hen than the Zhenning hen. Inspite of the similarity in the size of the oviduct between two breeds, Hyline hen had significantly higher oviduct weights. In addition to this, ovary weight and hierarchy was also bigger in the Hyline hen.Besides this, the histological structures were also showed remarkable difference between two breeds. Significant differences were observed between two breeds with respect to the egg quality parameters. White Leghorn hens had significantly higher egg weights and albumin weights in comparison with the Zhenning hens. Haugh unit and albumin height were also significantly higher in the Hyline hens than the Zhenning hens, In conclusion, these findings suggest that the Hyline and Zhenning hen differ significantly in many aspects terms of increase in body weight ratio of liver, ovary and oviduct morphology.2. Isolation, culture and morphological characterization of primordial folliclesThe success of the fertility in the female chickens depends on the successful development of the primordial follicles into the later stages by the regulating factors.The complete insight of these factors can only be determined by culturing the individual follicles which are tightly surrounded by the ovarian stroma. Thus, in our second experiment we have enzymatically isolated and cultured chicken primordial follicles by culturing them under different conditions. The objective of our work was to isolate healthy chicken primordial follicles in sufficient number and determine the appropriate culturing conditions required for their survival. For this purpose we have obtained the ovaries from the 2 week old chicken and have compared three different enzymes such as trypsin-EDTA, collagenase II and Collagenase IA, along with mechanical technique to isolate the chicken primordial follicles. Consequently, isolated follicles were counted and their viability was checked by the Neutral red stain. Later on the isolated follicles were cultured under different culturing conditions such as Alginate Hydrogel, Methylcellulose, Mineral oil drop, Floating water and on the Plane surface. The results showed that the maximum follicular yield was obtained by trypsin+EDTA and collagenase Ⅱ enzymes which is higher, then collagenase A and mechanical isolation method which shows a significant difference in the follicular recovery rate among different enzymes and methods used. Moreover, the highest follicular viability rate was observed in the mechanical isolated group and the collagenase A group. Additionally, enzymatic effect on the follicular morphology also differed among individual groups. Total number of misshapen oocytes was higher in collagenase II and collagenase IA whereas naked oocytes number was significantly higher in the trypsin+EDTA treated group. Besides, our follicle culture conditions results showed maximum survival rate of primordial follicle in the alginate hydrogel beads after 12 d culture. In addition, our morphological characterization results showed that the average diameter of primordial follicles ranged from 50 to 53 μm and the average number of flattened granulosa cells surrounding oocytes were 12 μm.Therefore, it was the first time that we have successfully isolated the chicken primordial follicles and cultured them; however more studies are required to work on the growth of the chicken primordial follicles.3. Effect of LIF on the growth of follicleIn our third experiment, we have investigated the effect of different factors on the growth of ovarian follicles at various stages.For this purpose, we selected LIF, and SCF, to observe their effect on the growth of the primordial, primary and small yellow follicles. We used in vitro techniques to study the effect of these factors on the growth of follicles. Cortical pieces approximately measuring lmm3 were obtained after the dissection of 16 d chicken ovary were cultured in the DMEM media and supported with ITS as supplement.LIF and SCF were added in different concentrations. BrdU incorporation and the histological methods were used to measure the growth of the follicles. Our histological and BrdU immunofluorescence results showed that LIF and SCF factors increased in the follicular growth in case of primordial follicle after 12 d of folhcular culture.Moreover, the BrdU incorporation also showed cellular proliferation in the primary and small yellow follicle walls during follicular culture after 24 h of follicle culture.Moreover, we have also confirmed the presence of LIF in the ovaries during the different stages of the development by using immunohistochemistry. Thus, on the basis of histological and immunohistochemical studies we observed that LIF and SCF were indirectly involved in growth of primordial, primary and small yellow follicles.In conclusion, we have compared the two different breeds of hens for their anatomical, morphological difference in the egg processing organs, and highlighted the major difference present in the follicular hierarchy, reproductive tract structures, and egg quality characteristics. We have also isolated, cultured and morphological characterized the chicken primordial follicles and found the most appropriate enzyme for the isolation of the primordial follicle and identified the culturing conditions required for the follicular survival and growth. We have also evaluated the LIF role in the follicular development at different stages. It is expected that the results will be helpful to the breeder companies and the researchers working on the different aspects of the follicular development in the in vitro studies. |
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