Effects Of Vitamin D Deficiency On Rats Lung Morphogenesis And Platelet-derived Growth Factor-a

Objective:Rat models were used to observe the lung morphogenesis and the expression of platelet-derived growth factor-A (PDGF-A), with the purpose making a further attempt to investigate the effects of vitamin D deficiency on lung morphogenesis and possible mechanism.Methods:Eighteen healthy nulliparous Sprague-Dawley female rats were divided into 3 groups randomly including normal control group, vitamin D deficiency experiment model group and intervention group, with 6 rats in each group. We fed the model and intervention groups with fodder without vitamin D away from light, while the control group received normal fodder and normal light for two weeks. Serum 25(OH)D of all the rats were detected by high-performance liquid chromatography (HPLC) before and after two weeks'different feeding respectively. With the verification that the vitamin D deficiency model was made successful, the female rats in 3 groups copulated with normal male rats. On days E17d, E18d and E19d, each rat of the intervention group received intragastric administration with active vitamin D (0.5mg/kg), and they regained light and normal fodder. Serum 25(OH)D of rats in each group were detected again through the method of HPLC. Cesarean sections were performed on 3 pregnant rats on the 20th day of gestation, while the other 3 rats waited till spontaneous delivery. The morphology of fetal rat lungs on E20d and the 1-day-old neonatal rat lungs was observed by light microscope and electronic microscope. RT-PCR, Western-Blot and immunohistochemistry methods were used respectively to detect the mRNA and protein level of PDGF-A gene.Results:1. No statistical difference of the original value of serum 25(OH)D was found among three groups(P>0.05). Two weeks later, however, the value of model group as well as intervention group was significantly lower than that of control group(P<0.05), suggesting vitamin D deficiency rat model made successfully. On E20d, serum 25(OH)D level of rat in intervention group was higher compared to model group, though still lower than control group (P<0.05).2. Under the light microscope, smaller alveolar space, smaller respiratory membrane's diameter and thicker alveolus mesenchyma was shown of vitamin D deficiency model group and intervention group compared with control group (P<0.05). Among those, larger alveolar space, larger respiratory membrane's diameter and thinner alveolus mesenchyma were seen in 1d neonatal rat lungs of intervention group while compared to experimental model group (P<0.05).3. With the electronic microscope, fewer lamellar bodies but more glycogen deposition in intracytoplasm was detected in the fetal lungs of generational age 20d in experimental model group than control. The above changes of lungs in intervention group were not as well as those in model group. On 1 day after birth, despite of number, the lamellar bodies were not as condensed in model group as in control group, what's more, empty lamellar bodies were found in model group. The evacuation phenomenon of lamellar bodies in type II epithelial cells was not seen in intervention group.4. RT-PCR results: The mRNA expression of PDGF-A gene in rat lung tissue was down-regulated in model group as well as intervention group compared to that of the control group (P<0.05). Besides, there existed statistical difference between intervention group and model group in levels of PDGF-A mRNA of 1d rat lungs (P<0.05).5. Immunohistochemistry results: PDGF-A was expressed in pulmonary bronchial and alveolar epithelium of both E20d fetal lungs and neonatal 1d rat lungs. The expression of PDGF-A in control group were much higher compared with vitamin D deficiency and intervention groups (P<0.05).6.Western-blot results: The expressions of PDGF-A protein in rat lungs of vitamin D deficiency group and intervention group were significantly lower than that of control group (P<0.05).Conclusions:1. Vitamin D deficiency in generation could inhibit the development of rat lung morphogenesis, with the manifestation of smaller alveolar space, smaller respiratory membrane's diameter and thicker alveolus mesenchyma, which could be improved by supplement of vitamin D.2. PDGF-A expresses in late fetal and newborn rat lungs. Vitamin D deficiency can inhibit the expression of PDGF-A mRNA as well as protein in rat lungs.3. The low expression of PDGF-A may be involved in the mechanism of effects on lung development by vitamin D deficiency.