Effect of Lactobacillus casei fermented milk on fracture healing in osteoporotic mice.

The interaction between the gut microbiota and the host has been described experimentally by germ-free animals or by antibiotic-disturbed gut microbiota. Studies on germ-free mice have shown that gut microbiota is critical for bone growth and development in mice, emphasizing that microbiota dysbiosis may interfere with normal bone development processes. This study aimed to clarify the effect of antibiotic treatment on disturbed gut microbiota on bone development in mice and to investigate the effect of probiotic treatment on fracture healing in mice with dysbiosis. Our results showed that 4 weeks old female Kunming mice showed significantly lower abundance and diversity of the gut microbiota and significantly lower bone mineral density after 12 weeks of antibiotic treatment and significantly increased levels of RANKL and Ang II in serum (p<0.05). Mice with dysbiosis received 5 mL of Lactobacillus casei fermented milk by daily gavage after internal fixation of femoral fractures, and postoperative fracture healing was evaluated by X-ray, micro-CT scan, and HE staining, which showed faster growth of the broken ends of the femur and the presence of more callus. Serological tests showed decreased levels of RANKL and Ang II (p<0.05). Similarly, immunohistochemical results also showed increased expression of α smooth muscle actin in callus tissue. These results suggest that oral antibiotics can lead to dysbiosis of the gut microbiota in mice, which in turn leads to the development of osteoporosis. In contrast, probiotic treatment promoted fracture healing in osteoporotic mice after dysbiosis, and the probiotic effect on fracture healing may be produced by inhibiting the RAS/RANKL/RANK pathway.

Gut microbiota can affect bone quality by regulating serum estrogen levels.

Germ-free (GF) animals and animal models of the antibiotic disruption of gut microbiota are widely used to explore studies of gut microbiota-host interactions. The role of gut microbiota in bone growth and development has been well explained in studies on GF mice, indicating that changes in the gut microbiota may affect normal bone developmental processes. The mechanisms, however, are yet unclear. This study aims to clarify the effect of antibiotic treatment disrupting the gut microbiota on bone development in mice and investigate the possible causes of this effect. Our results show that long-term antibiotic feeding significantly alters gut microbiota composition in mice, reduces the bone mineral density of the spinal region, and leads to changes in trabecular microstructure. Interestingly, we found a significant decrease in the serum estrogen levels in mice treated with antibiotics, suggesting that gut microbiota may affect bone quality by regulating serum estrogen levels. These results may help understand how gut ecological dysregulation affects sex hormones and provide a new conception for the clinical treatments of osteoporosis.

Adipose-derived mesenchymal stem cells with hypoxic preconditioning improve tenogenic differentiation.

BACKGROUND: Adipose-derived mesenchymal stem cells (ADSCs), as seed cells for tendon tissue engineering, are promising for tendon repair and regeneration. But for ADSCs, diverse oxygen tensions have different stimulatory effects. To explore this issue, we investigated the tenogenic differentiation capability of ADSCs under hypoxia condition (5% O2) and the possible signaling pathways correspondingly. The effects of different oxygen tensions on proliferation, migration, and tenogenic differentiation potential of ADSCs were investigated. METHODS: P4 ADSCs were divided into a hypoxic group and a normoxic group. The hypoxic group was incubated under a reduced O2 pressure (5% O2, 5% CO2, balanced N2). The normoxic group was cultured in 21% O2. Two groups were compared: HIF-1α inhibitor (2-MeOE2) in normoxic culturing conditions and hypoxic culturing conditions. Hypoxia-inducible factor-1α (HIF-1α) and VEGF were measured using RT-qPCR. Specific HIF-1α inhibitor 2-methoxyestradiol (2-MeOE2) was applied to investigate whether HIF-1α involved in ADSCs tenogenesis under hypoxia. RESULTS: Hypoxia significantly reduced proliferation and migration of ADSCs. Continuous treatment of ADSCs at 5% O2 resulted in a remarkable decrease in HIF-1α expression in comparison with 20% O2. Additionally, ADSCs of hypoxia preconditioning exhibited higher mRNA expression levels of the related key tenogenic makers and VEGF than normoxia via RT-qPCR measurement (p ˂ 0.05). Furthermore, the effects of hypoxia on tenogenic differentiation of ADSCs were inhibited by 2-MeOE2. Hypoxia can also stimulate VEGF production in ADSCs. CONCLUSIONS: Our findings demonstrate that hypoxia preconditioning attenuates the proliferation and migration ability of ADSCs, but has positive impact on tenogenic differentiation through HIF-1α signaling pathway.

Effects of hypoxia on Achilles tendon repair using adipose tissue-derived mesenchymal stem cells seeded small intestinal submucosa.

BACKGROUND: The study was performed to evaluate the feasibility of utilizing small intestinal submucosa (SIS) scaffolds seeded with adipose-derived mesenchymal stem cells (ADMSCs) for engineered tendon repairing rat Achilles tendon defects and to compare the effects of preconditioning treatments (hypoxic vs. normoxic) on the tendon healing. METHODS: Fifty SD rats were randomized into five groups. Group A received sham operation (blank control). In other groups, the Achilles tendon was resected and filled with the original tendon (Group B, autograft), cell-free SIS (Group C), or SIS seeded with ADMSCs preconditioned under normoxic conditions (Group D) or hypoxic conditions (Group E). Samples were collected 4 weeks after operation and analyzed by histology, immunohistochemistry, and tensile testing. RESULTS: Histologically, compared with Groups C and D, Group E showed a significant improvement in extracellular matrix production and a higher compactness of collagen fibers. Group E also exhibited a significantly higher peak tensile load than Groups D and C. Additionally, Group D had a significantly higher peak load than Group C. Immunohistochemically, Group E exhibited a significantly higher percentage of MKX + cells than Group D. The proportion of ADMSCs simultaneously positive for both MKX and CM-Dil observed from Group E was also greater than that in Group D. CONCLUSIONS: In this animal model, the engineered tendon grafts created by seeding ADMSCs on SIS were superior to cell-free SIS. The hypoxic precondition further improved the expression of tendon-related genes in the seeded cells and increased the rupture load after grafting in the Achilles tendon defects.

Wrist tuberculosis-experience from eighteen cases: a retrospective study.

BACKGROUND: Wrist tuberculosis is a rare disease, which is easy to be misdiagnosed, leading to delayed treatment and poor prognosis. In this study, the clinical manifestations, diagnosis, treatment, and prognosis of 18 cases of wrist tuberculosis were analyzed retrospectively. METHODS: A retrospective study was conducted, investigating tuberculosis of the wrist, diagnosed in 18 patients from August 2013 to November 2018. Puncture biopsy confirmed the diagnosis. The study includes 11 males and 7 females, and 8 left and 10 right wrists. The average age was 53.5 ± 18.3 years and ranged from 15 to 81 years. The disease course was 1 to 42 months, with an average of 15.1 ± 11.3 months. Eighteen patients were underwent surgery and chemotherapy, 3 patients with severe bone defects were treated with wrist fusion, and 15 patients were underwent focus removal. The Gartland and Werley score, DASH score, the range of motion (ROM), grip strength, and imaging examinations were used to evaluate the postoperative recovery of the patients. RESULTS: Eighteen patients were followed up for 15 to 77 months, with an average follow up of 39.7 ± 15.3 months. The ESR and CRP levels were normal for all patients after chemotherapy. No recurrence of tuberculosis was observed in any of the patients. Among the 15 focus removals, the Gartland and Werley scores at admission, two weeks of chemotherapy, 1 month after surgery, and the last follow-up were 21.73 ± 4.33, 18.60 ± 3.16,11.27 ± 2.79, and 5.07 ± 2.28, respectively; and the DASH scores were 45.87 ± 5.58, 39.47 ± 4.72, 22.67 ± 6.54, and 6.73 ± 2.94, respectively. The range of motion (ROM) of the wrist and grip strength improved significantly when compared to those at admission. Among the three cases of wrist fusion, 2 were fixed with a steel plate and the fixation position of wrist joint was good. One case was fixed with Kirschner wire and resulted in a slightly deformed wrist joint. CONCLUSION: For patients with wrist tuberculosis, early diagnosis, preoperative and postoperative chemotherapy, thorough focus removal, and appropriate fixation of the affected limb can help restore the function of the affected wrist, reduce the recurrence rate, and improve the quality of life.