Efficacy and mechanism of HO1 engineered cell-free extract combined with graphene far-infrared radiation in promoting the repair of diabetic ulcer
- JOURNAL OF REGIONAL ANATOMY AND OPERATIVE SURGERY Vol. 32, Issue 4, Pages: 285-292(2023)
- 作者机构：
1.陆军军医大学第一附属医院老年与特勤医学科，重庆 400038
2.陆军军医大学基础医学院组织学与胚胎学教研室/重庆市神经科学重点实验室，重庆 400038
3.陆军军医大学基础医学院基础医学教学实验中心，重庆 400038
- 作者简介：
- 基金信息：
DOI：10.11659/jjssx.01E023063
CLC： R587.2Published：25 April 2023，
Received：13 January 2023，
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温晴,范睿诚,杨艺等.HO1工程化无细胞提取液联合石墨烯远红外光促糖尿病溃疡修复的疗效及机制[J].局解手术学杂志,2023,32(04):285-292.
WEN Qing,FAN Rui-cheng,YANG Yi,et al.Efficacy and mechanism of HO1 engineered cell-free extract combined with graphene far-infrared radiation in promoting the repair of diabetic ulcer[J].JOURNAL OF REGIONAL ANATOMY AND OPERATIVE SURGERY,2023,32(04):285-292.
温晴,范睿诚,杨艺等.HO1工程化无细胞提取液联合石墨烯远红外光促糖尿病溃疡修复的疗效及机制[J].局解手术学杂志,2023,32(04):285-292. DOI： 10.11659/jjssx.01E023063.
WEN Qing,FAN Rui-cheng,YANG Yi,et al.Efficacy and mechanism of HO1 engineered cell-free extract combined with graphene far-infrared radiation in promoting the repair of diabetic ulcer[J].JOURNAL OF REGIONAL ANATOMY AND OPERATIVE SURGERY,2023,32(04):285-292. DOI： 10.11659/jjssx.01E023063.
摘要
目的
2
探讨HO1工程化无细胞提取液（CE）联合石墨烯远红外光（FIR）在糖尿病溃疡创面修复中的疗效及机制。
方法
2
将40只6~8周龄雄性C57BL/6小鼠建立正常创面模型（对照组，
n
=8）及糖尿病溃疡模型（
n
=32），其中将糖尿病溃疡模型小鼠随机分为糖尿病组、
HO1
BMSCs-CE组、FIR组、联合组。构建过表达血红素氧合酶1（HO1）的骨髓间充质干细胞（BMSCs），获取工程化富含HO1蛋白的无细胞提取液（
HO1
BMSCs-CE），运用Western blot、ELISA、RT-PCR、Masson染色、免疫组化等检测各组创面愈合率、组织修复情况、创面氧化应激及炎症水平等。
结果
2
Western blot及ELISA检测结果显示，
HO1
BMSCs-CE中HO1、表皮生长因子（EGF）、转化生长因子-β（TGF-β）表达水平显著高于BMSCs-CE（
P
<
0.05）。治疗后第5、11天，联合组的创面愈合率均显著高于
HO1
BMSCs-CE组和FIR组（
P
<
0.05）。与
HO1
BMSCs-CE组及FIR组相比，联合组在治疗后第3、7、11天创缘组织中Ca
2+
浓度较高，TNF-α、IL-6表达水平较低，而ROS水平仅在治疗后第3、7天较低（
P
<
0.05）。治疗后第14天，联合组创面组织的表皮厚度较厚，真皮Masson染色胶原容积分数和VEGF免疫阳性信号OD值均显著高于FIR组和
HO1
BMSCs-CE组（
P
<
0.05）；治疗后第7天，联合组创面组织中凋亡相关蛋白Caspase-3表达水平显著低于FIR组和
HO1
BMSCs-CE组（
P
<
0.05）。
结论
2
糖尿病溃疡创面中运用工程化富含HO1蛋白的
HO1
BMSCs-CE联合石墨烯FIR治疗可明显抑制局部组织氧化应激反应，减少炎症因子释放和细胞凋亡发生，加快创面区域皮肤组织上皮化形成、真皮胶原和血管增生，较
HO1
BMSCs-CE或FIR治疗的促愈疗效更为突出，为探索临床糖尿病难愈性创面的新型联合治疗提供了基础研究证据。
Abstract
Objective
2
To investigate the efficacy and mechanism of HO1 engineered cell-free extract（CE） combined with graphene far-infrared radiation （FIR） in the repair of diabetic ulcer wounds.
Methods
2
A total of 40 male C57BL/6 mice （6 to 8 weeks old） were used to establish normal wound model （the control group，
n
=8） and diabetic ulcer model （
n
=32）. The diabetic ulcer model mice were randomly divided into the diabetes group， the
HO1
BMSCs-CE group， the FIR group and the combined group. The bone marrow mesenchymal stem cells （BMSCs） overexpressing heme oxygenase-1（HO1） was constructed to obtain engineered cell-free extract rich in HO1 protein （
HO1
BMSCs-CE）. Western blot， ELISA， RT-PCR， Masson staining and immunohistochemistry were used to detect the wound healing rate， tissue healing status， wound oxidative stress and inflammation levels in each group.
Results
2
The results of Western blot and ELISA showed that the expression levels of HO1， epidermal growth factor （EGF） and transforming growth factor-β （TGF-β） in
HO1
BMSCs-CE were significantly higher than those in BMSCs-CE（
P
<
0.05）. The wound healing rate 5 and 11 days after treatment in the combined group were higher than those in the
HO1
BMSCs-CE group and the FIR group（
P
<
0.05）. Compared with the
HO1
BMSCs-CE group and the FIR group， the Ca
2+
concentration of the wound margin tissue 3， 7 and 11 days after treatment in the combined group were higher， and the expression levels of TNF-α and IL-6 were lower，while the expression levels of ROS 3 and 7 days after treatment were lower （
P
<
0.05）. The thickness of epidermis of the wound tissue 14 days after treatment in the combined group was thick， and the Masson staining collagen volume fraction in dermis and the OD value of VEGF immunopositive signal were significantly higher than those in the FIR group and the
HO1
BMSCs-CE group （
P
<
0.05）. The expression levels of apoptosis-related protein Caspase-3 of the wound tissue 7 days after treatment in the combined group was significantly lower than those in the FIR group and the
HO1
BMSCs-CE group （
P
<
0.05）.
Conclusion
2
The application of engineered
HO1
BMSCs-CE rich in HO1 protein combined with graphene FIR in diabetic ulcer wounds can significantly inhibit the oxidative stress response of local tissue， reduce the release of inflammatory factors and the occurrence of cell apoptosis， and accelerate the epithelization formation of skin tissue and the proliferation of dermal collagen and blood vessels in the wound area， which has more prominent curative effect than the efficacy of
HO1
BMSCs-CE or FIR therapy alone， and provide basic research evidence for the clinical exploration of new combined treatment for refractory diabetic wounds.
关键词
糖尿病溃疡骨髓间充质干细胞无细胞提取液血红素氧合酶1石墨烯远红外光
Keywords
diabetic ulcerbone marrow mesenchymal stem cellscell-free extractheme oxygenase 1graphene far-infrared radiation
references
蔡原，邓呈亮.脂肪干细胞无细胞提取液的治疗用途：皮肤老化、创面愈合、瘢痕恢复乃至神经再生［J］.中国组织工程研究，2021，25（13）：2097-2102.
He Y，Xia J，Chen H，et al.Human adipose liquid extract induces
angiogenesis and adipogenesis：a novel cell-free therapeutic agent［J］.Stem Cell Res Ther，2019，10（1）：252.doi：10.1186/s13287-019-1356-0http://dx.doi.org/10.1186/s13287-019-1356-0.
Feng W，Jin Q，Ming-Yu Y，et al.MiR-6924-5p-rich exosomes derived from genetically modified Scleraxis-overexpressing PDGFRalpha（+） BMMSCs as novel nanotherapeutics for treating osteolysis during tendon-bone healing and improving healing strength［J］.Biomaterials，2021，279：121242.doi：10.1016/j.biomaterials.2021.121242http://dx.doi.org/10.1016/j.biomaterials.2021.121242.
Jung B，Lee H，Kim S，et al.Effect of Rho-associated kinase inhibitor and mesenchymal stem cell-derived conditioned medium on corneal
endothelial cell senescence and proliferation［J］.Cells，2021，10（6）：1463.doi：10.3390/cells10061463http://dx.doi.org/10.3390/cells10061463.
L PK，Kandoi S，Misra R，et al.The mesenchymal stem cell secretome：a new paradigm towards cell-free therapeutic mode in regenerative medicine［J］.Cytokine Growth Factor Rev，2019，46：1-9.doi：10.1016/j.cytogfr.2019.04.002http://dx.doi.org/10.1016/j.cytogfr.2019.04.002.
Maamoun H，Zachariah M，Mcvey JH，et al.Heme oxygenase （HO）-1 induction prevents Endoplasmic Reticulum stress-mediated endothelial cell death and impaired angiogenic capacity［J］.Biochem Pharmacol，2017，127：46-59.doi：10.1016/j.bcp.2016.12.009http://dx.doi.org/10.1016/j.bcp.2016.12.009.
Wang YH，Cheng FY，Chao YC，et al.Effects of far-infrared therapy on foot circulation among hemodialysis patients with diabetes mellitus［J］.Biol Res Nurs，2020，22（3）：403-411.doi：10.1177/1099800420 923730http://dx.doi.org/10.1177/1099800420923730.
许祥生.石墨烯智能理疗护膝治疗膝关节骨性关节炎的疗效观察［J］.中医外治杂志，2020，29（1）：60-62.
Lin YH，Li TS.The application of far-infrared in the treatment of wound healing：a short evidence-based analysis［J］.J Evid Based Complementary Altern Med，2017，22（1）：186-188.doi：10.1177/2156587215623436http://dx.doi.org/10.1177/2156587215623436.
Ryzhii V，Ryzhii M，Mitin V，et al.Far-infrared photodetectors based on graphene/black-AsP heterostructures［J］.Opt Express，2020，
28（2）：2480-2498.doi：10.1364/OE.376299http://dx.doi.org/10.1364/OE.376299.
Lee DE，Ayoub N，Agrawal DK.Mesenchymal stem cells and cutaneous wound healing：novel methods to increase cell delivery and therapeutic efficacy［J］.Stem Cell Res Ther，2016，7：37.doi：10.1186/s13287-016-0303-6http://dx.doi.org/10.1186/s13287-016-0303-6.
苏龙辉，范少勇，双峰.慢性创面的治疗进展［J］.实用中西医结合临床，2020，20（14）：156-159.
Watanabe Y，Tsuchiya A，Terai S.The development of mesenchymal stem cell therapy in the present，and the perspective of cell-free therapy in the future［J］.Clin Mol Hepatol，2021，27（1）：70-80.doi：10.3350/cmh.2020.0194http://dx.doi.org/10.3350/cmh.2020.0194.
Cai Y，Li J，Jia C，et al.Therapeutic applications of adipose cell-free derivatives：a review［J］.Stem Cell Res Ther，2020，11（1）：312.doi：10.1186/s13287-020-01831-3http://dx.doi.org/10.1186/s13287-020-01831-3.
Chen QY，Wang GG，Li W，et al.Heme oxygenase-1 promotes
delayed wound healing in diabetic rats［J］.J Diabetes Res，2016，2016：9726503.doi：10.1155/2016/9726503http://dx.doi.org/10.1155/2016/9726503.
Grajek H，Jonik J，Witkiewicz Z，et al.Applications of graphene and its derivatives in chemical analysis［J］.Crit Rev Anal Chem，2020，
50（5）：445-471.doi：10.1080/10408347.2019.1653165http://dx.doi.org/10.1080/10408347.2019.1653165.
Zarei F，Negahdari B，Eatemadi A.Diabetic ulcer regeneration：stem cells，biomaterials，growth factors［J］.Artif Cells Nanomed Biotechnol，2018，46（1）：26-32.doi：10.1080/21691401.2017.1304407http://dx.doi.org/10.1080/21691401.2017.1304407.
Wilkinson HN，Hardman MJ.Wound healing：cellular mechanisms and pathological outcomes［J］.Open Biol，2020，10（9）：200223.doi：10.1098/rsob.200223http://dx.doi.org/10.1098/rsob.200223.
Pastore D，Pacifici F，Ciao G，et al.Far infrared technology （FIT） therapy patches，protects from inflammation， oxidative stress and
promotes cellular vitality［J］.Curr Pharm Des，2020，26（34）：4323-4329.doi：10.2174/1381612826666200427112023http://dx.doi.org/10.2174/1381612826666200427112023.
Ryter SW.Heme Oxgenase-1，a cardinal modulator of regulated cell death and inflammation［J］.Cells，2021，10（3）：515.doi：10.3390/cells10030515http://dx.doi.org/10.3390/cells10030515.
Korkmaz P，Kocak H，Onbasi K，et al.The role of serum procalcitonin，
interleukin-6，and fibrinogen levels in differential diagnosis of
diabetic foot ulcer infection［J］.J Diabetes Res，2018，2018：7104352.doi：10.1155/2018/7104352http://dx.doi.org/10.1155/2018/7104352.
Deng L，Du C，Song P，et al.The role of oxidative stress and antioxidants in diabetic wound healing［J］.Oxid Med Cell Longev，2021，2021：8852759.doi：10.1155/2021/8852759http://dx.doi.org/10.1155/2021/8852759.
Sun D，Cao H，Yang L，et al.MiR-200b in heme oxygenase-1-modified bone marrow mesenchymal stem cell-derived exosomes alleviates inflammatory injury of intestinal epithelial cells by targeting high
mobility group box 3［J］.Cell Death Dis，2020，11（6）：480.doi：10.1038/s41419-020-2685-8http://dx.doi.org/10.1038/s41419-020-2685-8.
Rege AS，Sudan DL.Autologous gastrointestinal reconstruction：
review of the optimal nontransplant surgical options for adults and children with short bowel syndrome［J］.Nutr Clin Pract，2013，28（1）：65-74.doi：10.1177/0884533612460405http://dx.doi.org/10.1177/0884533612460405.
Murray AR，Chen Q，Takahashi Y，et al.MicroRNA-200b downregulates oxidation resistance 1 （Oxr1） expression in the retina of type 1 diabetes model［J］.Invest Ophthalmol Vis Sci，2013，54（3）：1689-1697.doi：10.1167/iovs.12-10921http://dx.doi.org/10.1167/iovs.12-10921.
Bacci S.Cellular mechanisms and therapies in wound healing：looking toward the future［J］.Biomedicines，2021，9（11）：1611.doi：10.3390/biomedicines9111611http://dx.doi.org/10.3390/biomedicines9111611.
Subramaniam T，Fauzi MB，Lokanathan Y，et al.The role of calcium in wound healing［J］.Int J Mol Sci，2021，22（12）：6486.doi：10.3390/ijms22126486http://dx.doi.org/10.3390/ijms22126486.
Li S，Wang X，Chen J，et al.Calcium ion cross-linked sodium alginate hydrogels containing deferoxamine and copper nanoparticles for
diabetic wound healing［J］.Int J Biol Macromol，2022，202：657-670.doi：10.1016/j.ijbiomac.2022.01.080http://dx.doi.org/10.1016/j.ijbiomac.2022.01.080.
Navarro-Requena C，Perez-Amodio S，Castano O，et al.Wound healing-promoting effects stimulated by extracellular calcium and calcium-releasing nanoparticles on dermal fibroblasts［J］.Nanotechnology，2018，29（39）：395102.doi：10.1088/1361-6528/aad01fhttp://dx.doi.org/10.1088/1361-6528/aad01f.
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