Osteocytes and Primary Cilia.

PURPOSE OF REVIEW: The purpose of this review is to provide a background on osteocytes and the primary cilium, discussing the role it plays in osteocyte mechanosensing. RECENT FINDINGS: Osteocytes are thought to be the primary mechanosensing cells in bone tissue, regulating bone adaptation in response to exercise, with the primary cilium suggested to be a key mechanosensing mechanism in bone. More recent work has suggested that, rather than being direct mechanosensors themselves, primary cilia in bone may instead form a key chemo-signalling nexus for processing mechanoregulated signalling pathways. Recent evidence suggests that pharmacologically induced lengthening of the primary cilium in osteocytes may potentiate greater mechanotransduction, rather than greater mechanosensing. While more research is required to delineate the specific osteocyte mechanobiological molecular mechanisms governed by the primary cilium, it is clear from the literature that the primary cilium has significant potential as a therapeutic target to treat mechanoregulated bone diseases, such as osteoporosis.

Beneficial Effects of a Blended Fibroin/Aloe Gel Extract Film on the Biomolecular Mechanism(s) via the MAPK/ERK Pathway Relating to Diabetic Wound Healing.

In diabetic patients, the process of wound healing is usually delayed or impaired. A diabetic environment could be associated with dermal fibroblast dysfunction, reduced angiogenesis, the release of excessive proinflammatory cytokines, and senescence features. Alternative therapeutic treatments using natural products are highly demanded for their high potential of bioactive activity in skin repair. Two natural extracts were combined to develop fibroin/aloe gel wound dressing. Our previous studies revealed that the prepared film enhances the healing rate of diabetic foot ulcers (DFUs). Moreover, we aimed to explore its biological effects and underlying biomolecular mechanisms on normal dermal, diabetic dermal, and diabetic wound fibroblasts. Cell culture experiments showed that the γ-irradiated blended fibroin/aloe gel extract film promotes skin wound healing by enhancing cell proliferation and migration, vascular epidermal growth factor (VEGF) secretion, and cell senescence prevention. Its action was mainly linked to the activation of the mitogen-activated protein kinases/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway known to regulate various cellular activities, including proliferation. Therefore, the findings of this study confirm and support our previous data. The blended fibroin/aloe gel extract film displays a biological behavior with favorable properties for delayed wound healing and can be considered as a promising therapeutic approach in the treatment of diabetic nonhealing ulcers.

Evaluation of the safety and healing potential of a fibroin-aloe gel film for the treatment of diabetic foot ulcers.

OBJECTIVE: This study aimed to develop a wound dressing prepared from the blending of silkworm fibroin and aloe gel extract for use in the treatment of diabetic foot ulcers (DFUs). METHODS: Fibroin extracted from silkworm cocoons and aloe gel extract were dissolved in deionised water. pH levels were then adjusted with lactic acid solution. A simple casting technique was used to obtain the fibroin-aloe gel film. The surface morphology, hardness, flexibility and infrared spectrum of the sterilised film were tested. Swelling ratio was measured from changes in weight. The cytocompatibility of the film to human dermal fibroblast was determined using XTT assay. Hard-to-heal DFUs (grade I Wagner score) were treated with the film for four weeks. The application site was assessed for allergic reactions and/or sensitisation. Wound size was measured using standardised digital photography. RESULTS: A total of five hard-to-heal DFUs were treated. The obtained film sterilised with ozonation showed a non-porous structure. The elongation at break and tensile strength of the wet film were 9.00±0.95% and 6.89±1.21N, respectively. Fourier-transform infrared spectroscopy data indicated the presence of amides I, II and III, of peptide linkage, which are the chemical characteristics of the fibroin. Functional groups relating to healing activity of the aloe gel extract were also found. The swelling ratio of the film immersed in water for 24 hours was 0.8±0.01. In three DFUs (40-50mm2 in size), a wound area reduction of 0.4-0.8mm2/day was observed and were healed in 2-3 weeks. The remaining two SFUs (500mm2 in size) showed a wound area reduction of 4mm2/day and were almost closed at four weeks. No allergic reaction or infection was observed in any of the wounds. CONCLUSION: The obtained film showed a non-porous structure, and its strength and flexibility were adequate for storage and handling. The film tended to increase the proliferation of fibroblasts. The wound dressing showed potential for accelerating the healing rate of DFUs.

Primary cilia respond to fluid shear stress and mediate flow-induced calcium deposition in osteoblasts.

Bone turnover in vivo is regulated by mechanical forces such as shear stress originating from interstitial oscillatory fluid flow (OFF), and bone cells in vitro respond to mechanical loading. However, the mechanisms by which bone cells sense mechanical forces, resulting in increased mineral deposition, are not well understood. The aim of this study was to investigate the role of the primary cilium in mechanosensing by osteoblasts. MLO-A5 murine osteoblasts were cultured in monolayer and subjected to two different OFF regimens: 5 short (2 h daily) bouts of OFF followed by morphological analysis of primary cilia; or exposure to chloral hydrate to damage or remove primary cilia and 2 short bouts (2 h on consecutive days) of OFF. Primary cilia were shorter and there were fewer cilia per cell after exposure to periods of OFF compared with static controls. Damage or removal of primary cilia inhibited OFF-induced PGE2 release into the medium and mineral deposition, assayed by Alizarin red staining. We conclude that primary cilia are important mediators of OFF-induced mineral deposition, which has relevance for the design of bone tissue engineering strategies and may inform clinical treatments of bone disorders causes by load-deficiency.

Effects of the blended fibroin/aloe gel film on wound healing in streptozotocin-induced diabetic rats.

Delayed healing remains a major clinical problem and here we have sought to develop an improved dressing film comprising 1.95% w/v fibroin and 0.05% w/v aloe gel extract. The tensile strength of dry film was 21.1 ± 0.5 MPa and broke at 1.1 ± 0.2% elongation; corresponding values for wet film were 18.3 ± 1.3 MPa and 1.9 ± 0.1%. The film maintained its shape upon water immersion and the swelling ratio of the dry film was 0.8 ± 0.1 while the water uptake was 43.7 ± 2.6%. After 28 days of incubation in phosphate buffered saline (1 M, pH 7.4, 37 °C), the weight of film was reduced by 6.7 ± 1.1% and the tensile strength and elongation at breaking point (dry state) were 15.4 ± 0.6 MPa and 1.5 ± 0.2%, respectively. Compared to aloe-free fibroin film (2.0% fibroin extract only), the blended film enhanced the attachment and proliferation of skin fibroblasts. The bFGF immunofluorescence of fibroblasts cultured on the blended film appeared greater than those cultured on tissue culture plate or on aloe-free fibroin film while α-smooth muscle actin was maintained. In streptozotocin-induced diabetic rats, the wounds dressed with the blended film were smaller (p <0.05) by day 7 after wounding, compared to untreated diabetic wounds. Histology of repaired diabetic wounds showed the fibroblast distribution and collagen fiber organization to be similar to wounds in normal rats, and this was matched by enhanced hydroxyproline content. Thus, such accelerated wound healing by the blended fibroin/aloe gel films may find application in treatment of diabetic non-healing skin ulcers.