Combined effects of low temperature, hyperosmolarity and seawater-conditioned pathogens on open fracture healing in a rat model simulating circumpolar environments

Oct 9, 2025Journal of orthopaedic surgery and research

How Cold, Salty Conditions, and Seawater Germs Together Affect Open Bone Fracture Healing in Rats Simulating Arctic Environments

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Abstract

Fracture healing was significantly impaired in rats exposed to circumpolar seawater immersion compared to those without exposure.

X-ray, micro-CT, and histological analyses indicated a delay in fracture healing for the seawater group.
Bacterial colony counts on internal fixation devices were highest in the seawater immersion group.
Leukocyte and neutrophil counts were significantly elevated on days 3 and 7 in the seawater group.
Serum levels of alkaline phosphatase (ALP) and vascular endothelial growth factor (VEGF) were significantly reduced in the seawater group.
The rates of bone union and callus maturation were significantly lower in the seawater group compared to the other treatment conditions.
Metagenomic analysis identified Flavobacterium, Rhodobacter, and Bacteroides as the dominant pathogens present in seawater.

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OBJECTIVE: To investigate the factors influencing the healing of in circumpolar latitude region seawater immersion conditions.

MATERIALS AND METHODS: A femoral fracture model was established in ninety 6-to-8-week-old male Sprague-Dawley rats, randomly assigned to five groups (n = 18 per group): (1) fracture only, (2) fracture with circumpolar seawater immersion, (3) fracture with low-temperature isotonic solution immersion, (4) fracture with aseptic circumpolar seawater immersion, and (5) fracture with low-temperature aseptic circumpolar seawater immersion. Fractures were confirmed postoperatively by radiographs on days 7, 21, and 42. Micro-CT and H&E staining were performed on day 42 to assess bone healing. Bacterial cultures from internal fixation devices were analyzed on day 3. Blood samples were collected on days 3, 7, and 14 to assess leukocyte and neutrophil counts, and serum ALP and VEGF levels were measured on days 7, 14, and 21. Pathogenic microorganisms in the seawater were identified by metagenomic analysis. Fracture healing and callus formation rates were compared using the Log-rank test.

RESULTS: X-ray, micro-CT, and histological analyses revealed significantly impaired fracture healing in the group exposed to circumpolar seawater immersion compared to the fracture-only group (P < 0.05). Bacterial colony counts on internal fixation devices were highest in the circumpolar seawater group (P < 0.05). Leukocyte and neutrophil levels were significantly elevated in this group on days 3 and 7 (P < 0.05), with no significant differences observed on day 14 (P > 0.05). Serum ALP and VEGF levels were significantly reduced on days 7, 14, and 21 (P < 0.05), although ALP levels on day 21 showed no significant difference (P > 0.05). Log-rank analysis indicated that the bone union and callus maturation rates were significantly lower in the circumpolar seawater group compared to the other four groups. Metagenomic analysis identified Flavobacterium, Rhodobacter, and Bacteroides as the dominant pathogens in circumpolar seawater.

CONCLUSIONS: This study demonstrates that , low temperature, and exposure to opportunistic pathogens under circumpolar seawater conditions collectively delay open fracture healing. Among these factors, opportunistic pathogens exert the most significant impact, highlighting microbial contamination as the primary barrier to bone regeneration in such environments and providing direction for future therapeutic strategies.

CLINICAL TRIAL NUMBER: not applicable.

Key numbers

42%

Radiographic Union Rate

Compared to 85% in the control group.

highest in seawater group

Bacterial Colony Counts

Bacterial counts were significantly higher in the seawater group.

significantly lower in seawater group

Serum ALP and VEGF Levels

Measured during the first two postoperative weeks.

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Combined effects of low temperature, hyperosmolarity and seawater-conditioned pathogens on open fracture healing in a rat model simulating circumpolar environments

Abstract

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