Histological and contractile changes in the genioglossus muscle after nasal obstruction in growing rats

Abstract

The aim of the study was to address the genioglossus muscle physiological and histological changes after unilateral nasal obstruction in growing rats. Fifty-four 6-day-old male Wistar albino rats were randomly divided into control (n = 27) and experimental (n = 27) groups. Unilateral nasal obstruction was performed at 8 days old. Contractile properties of the genioglossus whole muscle were measured at 5-, 7- and 9-week-old, including the twitch and tetanic forces, contraction time, half-decay time, and fatigue index. The histological characteristics of the genioglossus were also evaluated at 5-, 7- and 9-week-old, analyzing the myosin heavy chain composition of the slow, fast, IIa and IIb muscle fiber type, by measuring the number, rate, diameter and cross-sectional area. The maximal twitch force, and tetanic force at 60 Hz and 80 Hz force was significantly increased at all ages after nasal obstruction. The fatigue index was decreased at 5 weeks-old after nasal obstruction. The diameter and cross-sectional area of the fast, IIa and IIb muscle fiber types were increased at 7 and 9 weeks after nasal obstruction, while only the diameter of IIa type and cross-sectional area of IIb type were increased at 5 weeks-old after nasal obstruction. Nasal obstruction during growth affects the whole genioglossus muscle contractile properties and histological characteristics, increasing its force, the diameter and area of its muscle fibers. These changes in the genioglossus muscle may affect the normal growth, development and function of the craniofacial complex.

Figure 1

Organigram of experiment procedures. A sham procedure was performed at 8 days old in the control group, while a unilateral nasal obstruction by cauterization of the left nostril was performed in the experimental (nose obstruction) group at 8 days old. Data collection for the contractile properties of the genioglossus muscle, and histological analysis was performed at 5-, 7- and 9-week-old animals.

Figure 2

Contractile properties of Genioglossus muscle. (A) Maximal twitch force in grams (g) at 5, 7 and 9 weeks old. (B) Contraction time at 5, 7 and 9 weeks old. (C) Half-decay time at 5, 7 and 9 weeks old. (D) Tetanic force at 60 Hz at 5, 7 and 9 weeks old. (E) Tetanic force at 80 Hz at 5, 7 and 9 weeks old. (F) Fatigue index at 5, 7 and 9 weeks old. Bars in black, Control group; bars in grey, nose obstruction group. The x-axis shows the age of the rats, the y-axis shows the contractile properties measurements, for both control and nose obstruction groups. A repeated-measures multivariate analysis of variance was performed for intergroup and intragroup statistical comparisons. A simple main-effects analysis with Sidak adjustment was performed for multiple comparisons. Data are expressed as mean ± standard deviation. *^,#P < 0.05. Error bars at 95% confidence interval. 5 5 weeks old, 7 7 weeks old, 9 9 weeks old, g grams, ms millisecond, % percentage.

Figure 3

Diameter of the genioglossus muscle fibers. (A) Diameter of the slow-type muscle fibers at 5, 7 and 9 weeks old. (B) Diameter of the fast-type muscle fibers at 5, 7 and 9 weeks old. (C) Diameter of the fast IIa-type muscle fibers at 5, 7 and 9 weeks old. (D) Diameter of the fast IIb-type muscle fibers at 5, 7 and 9 weeks old. Bars in black, Control group; bars in grey, nose obstruction group. The x-axis shows the age of the rats, the y-axis shows the diameter measurement of muscle fibers, for both control and nose obstruction groups. A repeated-measures multivariate analysis of variance was performed for intergroup and intragroup statistical comparisons. A simple main-effects analysis with Sidak adjustment was performed for multiple comparisons. Data are expressed as mean ± standard deviation. *^,#P < 0.05. Error bars at 95% confidence interval. MHC-slow Myosin heavy chain expression of slow-type muscle fiber, MHC-fast Myosin heavy chain expression of fast-type muscle fiber, MHC-IIa Myosin heavy chain expression of IIa fast-type muscle fiber, MHC-IIb Myosin heavy chain expression of IIb fast-type muscle fiber, 5 5 weeks old, 7 7 weeks old, 9 9 weeks old, g grams, ms millisecond, % percentage.

Figure 4

Cross-sectional area of the genioglossus muscle fibers. (A) Cross-sectional area of the slow-type muscle fibers at 5, 7 and 9 weeks old. (B) Cross-sectional area of the fast-type muscle fibers at 5, 7 and 9 weeks old. (C) Cross-sectional area of the fast IIa-type muscle fibers at 5, 7 and 9 weeks old. (D) Cross-sectional area of the fast IIb-type muscle fibers at 5, 7 and 9 weeks old. Bars in black, Control group; bars in grey, nose obstruction group. The x-axis shows the age of the rats, the y-axis shows the cross-sectional measurement of muscle fibers, for both control and nose obstruction groups. A repeated-measures multivariate analysis of variance was performed for intergroup and intragroup statistical comparisons. A simple main-effects analysis with Sidak adjustment was performed for multiple comparisons. Data are expressed as mean ± standard deviation. *^,#P < 0.05. Error bars at 95% confidence interval. MHC-slow Myosin heavy chain expression of slow-type muscle fiber, MHC-fast Myosin heavy chain expression of fast-type muscle fiber, MHC-IIa Myosin heavy chain expression of IIa fast-type muscle fiber, MHC-IIb Myosin heavy chain expression of IIb fast-type muscle fiber, 5 5 weeks-old, 7 7 weeks-old, 9 9 weeks-old, g grams, ms millisecond, % percentage.

Figure 5

Representative images of the genioglossus fast-type muscle fibers in the 5-, 7-, and 9-week-old rats. Figures at ×20 magnitude. 1000 μm scale bar. MHC-fast Myosin heavy chain expression of fast-type muscle fiber.