目的:阴道分娩是盆底肌肉损伤和功能障碍的关键危险因素,以及随后的盆底疾病。多重平价进一步加剧了这些风险。使用大鼠模型,对人体盆底肌肉的研究进行了验证,我们之前发现,一次模拟出生损伤会导致盆底肌肉萎缩和纤维化.我们假设多胎损伤会进一步压倒肌肉再生能力,导致长期功能相关的病理改变。
方法:Sprague-Dawley大鼠经历了模拟出生损伤,并在经历额外的出生损伤之前允许其恢复8周。在急性(损伤后3天和7天)处死动物,亚急性(21-,28-,和受伤后35天),和长期(受伤后8周和12周)第二次受伤后的时间点(N=3-8/时间点),收集大鼠肛提肌复合体的耻骨尾肌部分,以评估反复出生损伤对肌肉机械和组织形态学特性的影响。通过定制的NanoString面板评估伴随的转录变化。未受伤的动物用作对照。通过双向方差分析对具有参数分布的数据进行分析,然后使用Tukey's或Sidak's检验进行事后配对比较;将非参数分布的数据与Kruskal-Wallis检验进行比较,然后与Dunn检验进行配对比较。数据,使用GraphPadPrismv8.0,圣地亚哥,CA,以平均值±SEM或中值(范围)表示。
结果:在第一次模拟出生伤害之后,相对于未受伤的对照组,主动肌力急剧下降(12.9±0.9vs25.98±2.1g/mm2,P<0.01)。4周时,在出生损伤后8周,肌肉活动力的产生恢复到基线水平,且无变化(P>0.99).同样,反复出生后立即观察到主动力急剧下降(18.07±1.2vs25.98±2.1g/mm2,P<0.05)。与一次出生损伤后的功能恢复相反,反复出生损伤后12周肌肉收缩功能长期下降(18.3±1.6vs25.98±2.1g/mm2,P<0.05)。与未受伤组相比,第二次受伤后的长期时间点纤维尺寸较小(12周vs未受伤对照组:1485(60.7-5000)vs1989(65.6-4702)μm2,P<0.0001)。具有中心核的纤维的比例,表明活跃的肌纤维再生,在第1次出生损伤后8周恢复到基线,(P=0.95),但在第2次损伤后12周仍保持升高(12周vs未受伤对照组:7.1±1.5vs0.84±0.13%,P<0.0001)。与第1次损伤后4周后肌肉内胶原蛋白含量稳定相反,纤维化变性在反复损伤后的12周内逐渐增加(12周vs未受伤的对照组:6。7±1.1vs2.03±0.2%,P<0.001)。与单次出生损伤相比,重复后观察到促炎基因的长时间表达伴随着更多的免疫浸润。
结论:总体而言,与单一伤害相比,重复的出生伤害导致更严重的病理改变,在大鼠模型中导致更明显的盆底肌肉变性和肌肉功能障碍。以上提供了多胎和女性盆底功能障碍风险增加之间的推定机械联系。
OBJECTIVE: Vaginal childbirth is a key risk factor for pelvic floor muscle injury and dysfunction, and subsequent pelvic floor disorders.
Multiparity further exacerbates these risks. Using the rat model, validated for the studies of the human pelvic floor muscles, we have previously identified that a single simulated birth injury results in pelvic floor muscle atrophy and fibrosis. We hypothesized that multiple birth injuries would further overwhelm the muscle regenerative capacity, leading to functionally relevant pathological alterations long-term.
METHODS: Sprague-Dawley rats underwent simulated birth injury and were allowed to recover for 8 weeks before undergoing additional birth injury. Animals were sacrificed at acute (3- and 7-days post injury), subacute (21-, 28-, and 35-days post-injury), and long-term (8- and 12-weeks post-injury) time points post-second injury (N=3-8/time point), and the pubocaudalis portion of the rat levator ani complex was harvested to assess the impact of repeated birth injuries on muscle mechanical and histomorphological properties. The accompanying transcriptional changes were assessed by a customized NanoString panel. Uninjured animals were used as controls. Data with a parametric distribution were analyzed by a two-way analysis of variance followed by post hoc pairwise comparisons using Tukey\'s or Sidak\'s tests; non-parametrically distributed data were compared with Kruskal-Wallis test followed by pairwise comparisons with Dunn\'s test. Data, analyzed using GraphPad Prism v8.0, San Diego, CA, are presented as mean ± SEM or median (range).
RESULTS: Following the 1st simulated birth injury, active muscle force decreased acutely relative to uninjured controls (12.9±0.9 vs 25.98±2.1 g/mm2, P<0.01). At 4 weeks, muscle active force production recovered to baseline and remained unchanged at 8 weeks after birth injury (P>0.99). Similarly, precipitous decrease in active force was observed immediately after repeated birth injury (18.07±1.2 vs 25.98±2.1 g/mm2, P<0.05). In contrast to the functional recovery after a single birth injury, a long-term decrease in muscle contractile function was observed up to 12 weeks after repeated birth injuries (18.3±1.6 vs 25.98±2.1 g/mm2, P<0.05). Fiber size was smaller at the long-term time points after 2nd injury compared to the uninjured group (12 weeks vs uninjured control: 1485 (60.7-5000) vs 1989 (65.6-4702) μm2, P<0.0001). The proportion of fibers with centralized nuclei, indicating active myofiber regeneration, returned to baseline at 8 weeks post-1st birth injury, (P=0.95), but remained elevated as far as 12 weeks post-2nd injury (12 weeks vs uninjured control: 7.1±1.5 vs 0.84±0.13%, P<0.0001). In contrast to the plateauing intramuscular collagen content after 4 weeks post-1st injury, fibrotic degeneration increased progressively over 12 weeks after repeated injury (12 weeks vs uninjured control: 6. 7±1.1 vs 2.03±0.2%, P<0.001). Prolonged expression of pro-inflammatory genes accompanied by a greater immune infiltrate was observed after repeated compared to a single birth injury.
CONCLUSIONS: Overall, repeated birth injuries led to a greater magnitude of pathological alterations compared to a single injury, resulting in more pronounced pelvic floor muscle degeneration and muscle dysfunction in the rat model. The above provides a putative mechanistic link between
multiparity and the increased risk of pelvic floor dysfunction in women.