Scientia Agricultura Sinica ›› 2023, Vol. 56 ›› Issue (11): 2202-2211.doi: 10.3864/j.issn.0578-1752.2023.11.013

• ANIMAL SCIENCE·VETERINARY SCIENCE • Previous Articles     Next Articles

The Micro-Structure of Tibetan Sheep Lung and Its HIF-1α and AQP1 Expression Characteristics

AYIMUGULI Abudureyimu1(), ZHANG Chen1, CAI Yong2, QIN Sheng1, LUO WenXue3, ZHAXIYINGPAI1   

  1. 1 College of Life Science and Engineering, Northwest Minzu University, Lanzhou 730030
    2 Department of Experimental Teaching, Northwest Minzu University, Lanzhou 730030
    3 Tianzhu Tibetan Autonomous County Animal Husbandry Technology Station, Wuwei 733200, Gansu
  • Received:2021-12-22 Accepted:2023-03-23 Online:2023-06-01 Published:2023-06-19

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Abstract:

【Background】HIF-1α is one of the key factors for cells to make adaptive response to hypoxia stress. It mainly maintains the balance of oxygen supply by regulating gene transcription. Aquaporins (AQPs) are hydrophobic transmembrane transporters regulating water homeostasis. Among them, AQP-1 mainly regulates the water transport among alveoli, pulmonary interstitium and capillaries, so as to maintain the fluid balance in the lung. The plateau environment is characterized by low oxygen, extremely cold, strong wind and radiation, so there are relatively few species which could adapt and survive. As the unique sheep species adapt to high altitude and high cold climate, Tibetan sheep has formed a special morphological structure and physiological functions adapted to the plateau environment. As the main executive organ of respiration, lung is very sensitive to hypoxia. 【Objective】 This study aimed to explore the structural characteristics of Tibetan sheep lung and the expression characteristics of HIF-1α and AQP1, so as to reveal the related roles of HIF-1α and AQP1 in Tibetan sheep high altitude adaption. 【Method】 The histochemical HE, PAS and Masson staining, immuno-histochemical SP and real-time fluorescence quantitative were used.【Result】The tunica thickness of lung in Tibetan sheep was 40.28 μm, which showed no significant difference with that of Small Tail Han sheep, but the elastic fiber proportion was significantly higher than that of Small Tail Han sheep (P<0.05); the number of goblet cells in bronchiolar epithelium in Tibetan sheep was significantly more than that in Small Tail Han sheep (P<0.05), but there was no significant difference in superior branches; some goblet cells were still detected in the epithelium of bronchioli terminales of Tibetan sheep; the thickness of smooth muscle of bronchioli terminales of Tibetan sheep was significantly thicker than that of Small Tail Han sheep (P<0.05); the proportion of smooth muscle in the pulmonary arteriole (diameter less than 100 μm) in Tibetan sheep was significantly less than that of Small Tail Han sheep; the thickness of respiratory bronchiole smooth muscle and the number of capillaries in Tibetan sheep were significantly higher than those in Small Tail Han sheep. The HIF-1α protein was mainly expressed in bronchiolar epithelium, pulmonary micro-vascular endothelium and alveolar septum, and mainly detected in cytoplasm; both Tibetan sheep and Small Tail Han sheep, the expression of HIF-1α was significantly higher in high altitude than that of low altitude, and HIF-1α expression in low altitude Tibetan sheep was significantly stronger than that of Small Tail Han sheep living at low altitude. AQP1 protein was mainly expressed in alveolar epithelium, alveolar septum, pulmonary microvascular endothelium, submucosal and smooth muscle of bronchioles, mainly located on cell membrane; the expression of AQP1 in lung of Tibetan sheep living at high altitude was significantly stronger than that of Tibetan sheep from low altitude, and also stronger than that of Small Tail Han sheep either altitude (P<0.05), but there was no significant difference between Small Tail Han sheep living at high altitude and low (P>0.05).【Conclusion】All those results indicated that the lung capsule of Tibetan sheep contained more elastic fibers and less microvascular smooth muscle than that of Small Tail Han sheep, with more number of goblet cells and smooth muscle in respiratory bronchioles. The expression of HIF-1α and AQP1 in Tibetan Sheep lung were significantly stronger than that of Small Tail Han sheep, and their expression increased with living altitude increasing.

Key words: Tibetan sheep, HIF-1α, AQP1, Lung

Table 1

Primer sequence"

引物名称
Primer name		 序列号
Accession No.		 序列
Sequence (5′-3′)		 扩增长度
Amplicon size (bp)		 退火温度
Tm (℃)
绵羊 β-Actin
Sheep β-Actin		 NM_001009784.3 F: GCAGATGTGGATCAGCAAGC
R:TCTCGTTTTCTGCGCAAGTT		 133 60
绵羊HIF-1α
Sheep HIF-1α		 XM_027971915.2 F: ATGTCTCCATTACCTGCCTCTGA
R:CTGTTAGGCTCAGGTGAACTTTGT		 197 56
绵羊AQP
Sheep AQP		 NM_001009194.1 F: ATCGTCGCCACTGTCATCCT
R:TATCGCCAGCAGGTGTCCAA		 240 58

Fig. 1

The micro-structure of the Tibetan sheep lung PM: Tunica; LA: Pulmonary arterioles; TB: Terminal bronchioles; HC: Hyaline cartilage; MG: Mixed glands；AD: Alveolar duct; AS: Alveolar sac; PA: Alveolar. A: Pulmonary capsule (Gomori×100); B: Elastic fibers (Gomori×100); C: Collagenous fibers (Masson×200); D: Bronchiole (H.E×40); E: Bronchioli terminals (H.E×100); F: Bronchial mucosa and glands (PAS×200); G: Goblet cells (PAS×400); H: Mixed Glands (HE×400); I: Respiratory bronchiole (Masson×200). J: Alveolar duct; K: Alveolar sac and alveolar (H.E×400)"

Table 2

The thickness and elastic fiber ratio in lung tunica in the Tibetan sheep and Small Tail Han sheep"

浆膜厚度
Tunica thickness (μm)		 弹性纤维厚度
Elastic fibers thickness (μm)		 弹性纤维百分比
Thickness proportion (%)
藏绵羊Tibetan sheep 40.28±1.58ns 21.95±1.65* 54.49*
小尾寒羊Small Tail Han sheep 39.24±2.15 12.25±1.55 31.21

Table 3

Differences of respiratory parts of lung in the Tibetan sheep and Small Tail Han sheep"

Reb平滑肌厚度
Smooth muscle of Reb (μm)		 单个肺泡面积
APA (μm2)		 肺泡隔毛细血管数
Capillary number (mm2)
藏绵羊 Tibetan sheep 25.12±1.25* 1256±56ns 113±12*
小尾寒羊 Small Tail Han sheep 8.05±0.75 1368±42 65±9

Table 4

The ratio of smooth muscle layer to pulmonary arterioles in the Tibetan sheep and Small Tail Han sheep (%)"

肺微动脉直径 Pulmonary arteriole diameter (μm)
<50 50—100 101—200 201—400
藏绵羊 Tibetan sheep 28.14±3.14a 22.32±5.14a 20.12±1.99a 18.24±3.21a
小尾寒羊 Small Tail Han sheep 36.36±2.36b 34.25±2.14b 21.24±2.24a 18.66±4.56a

Table 5

The differences in goblet cell numbers and smooth muscle thickness in Bronchioli of the Tibetan sheep and Small Tail Han sheep"

杯状细胞数Goblet cell numbers (mm2) 平滑肌厚度Smooth muscle thickness (μm)
藏绵羊
Tibetan sheep		 小尾寒羊
Small Tail Han sheep		 藏绵羊
Tibetan sheep		 小尾寒羊
Small Tail Han sheep
段支气管 Segmental bronchi 1854±98a 1759±65a 215.12±12.24a 204.26±14.35a
小支气管 Bronchium 1453±54a 1398±62a 175.14±4.13a 168.24±3.65a
细支气管 Bronchiole 541±24b 142±12c 45.34±2.55b 42.24±3.65b
终末细支气管 Bronchioli terminales 15±3d ND 36.14±1.25b 21.09±2.34c

Fig. 2

Expression of HIF-1α and AQP1 protein in lung of the Tibetan sheep and Small Tail Han sheep（×100） a: HIF-1α expression in the lung of Tibetan sheep in higher altitude; b: HIF-1α expression in the lung of Tibetan sheep in lower altitude; c: HIF-1α expression in the lung of Small Tail Han sheep in higher altitude; d: HIF-1α expression in the lung of Small Tail Han sheep in lower altitude; e: AQP1 expression in the lung of Tibetan sheep in higher altitude; f: AQP1 expression in the lung of Tibetan sheep in lower altitude; g: AQP1 expression in the lung of Small Tail Han sheep in higher altitude; h: AQP1 expression in the lung of Small Tail Han sheep in lower altitude"

Fig. 3

Expression of HIF-1α and AQP1 in lung of the Tibetan sheep and Small Tail Han sheep Different letters mean significant difference (P<0.05), and same letters mean no significant difference (P>0.05). TH: Tibetan sheep in higher altitude; TL: Tibetan sheep in lower altitude; SH: Small Tail Han sheep in higher altitude; SL: Small Tail Han sheep in lower altitude"

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