Identification and validation of NAD+ metabolism-related biomarkers in patients with diabetic peripheral neuropathy.

Background: The mechanism of Nicotinamide Adenine Dinucleotide (NAD+) metabolism-related genes (NMRGs) in diabetic peripheral neuropathy (DPN) is unclear. This study aimed to find new NMRGs biomarkers in DPN. Methods: DPN related datasets GSE95849 and GSE185011 were acquired from the Gene Expression Omnibus (GEO) database. 51 NMRGs were collected from a previous article. To explore NMRGs expression in DPN and control samples, differential expression analysis was completed in GSE95849 to obtain differentially expressed genes (DEGs), and the intersection of DEGs and NMRGs was regarded as DE-NMRGs. Next, a protein-protein interaction (PPI) network based on DE-NMRGs was constructed and biomarkers were screened by eight algorithms. Additionally, Gene Set Enrichment Analysis (GSEA) enrichment analysis was completed, biomarker-based column line graphs were constructed, lncRNA-miRNA-mRNA and competing endogenouse (ce) RNA networks were constructed, and drug prediction was completed. Finally, biomarkers expression validation was completed in GSE95849 and GSE185011. Results: 5217 DEGs were obtained from GSE95849 and 21 overlapping genes of DEGs and NMRGs were DE-NMRGs. Functional enrichment analysis revealed that DE-NMRGs were associated with glycosyl compound metabolic process. The PPI network contained 93 protein-interaction pairs and 21 nodes, with strong interactions between NMNAT1 and NAMPT, NADK and NMNAT3, ENPP3 and NUDT12 as biomarkers based on 8 algorithms. Expression validation suggested that ENPP3 and NUDT12 were upregulated in DPN samples (P < 0.05). Moreover, an alignment diagram with good diagnostic efficacy based on ENPP3 and NUDT12 were identified was constructed. GSEA suggested that ENPP3 was enriched in Toll like receptor (TLR) pathway, NUDT12 was enriched in maturity onset diabetes of the young and insulin pathway. Furthermore, 18 potential miRNAs and 36 Transcription factors (TFs) were predicted and the miRNA-mRNA-TF networks were constructed, suggesting that ENPP3 might regulate hsa-miR-34a-5p by affecting MYNN. The ceRNA network suggested that XLOC_013024 might regulate hsa-let-7b-5p by affecting NUDT12. 15 drugs were predicted, with 8 drugs affecting NUDT12 such as resveratrol, and 13 drugs affecting ENPP3 such as troglitazone. Conclusion: ENPP3 and NUDT12 might play key roles in DPN, which provides reference for further research on DPN.

Free Chimeric Superficial Circumflex Iliac Artery Perforator Flap in Reconstructing the Distal Complex Extensor Tendon Injury.

SUMMARY: The distal complex extensor tendon injury, presenting as traumatic skin, zones 1 and 2 of extensor pollicis longus and extensor hallucis longus, and bony insertion loss, represents a challenging issue and requires a well-vascularized skin paddle, tendinous graft, and insertional reconstruction. Guided by the all-in-one-step reconstruction rule, the chimeric superficial circumflex iliac artery perforator (SCIAP) flap, generally considered as a promising multiple-type tissue provider (eg, vascularized skin paddle, fascia, iliac flap), can fulfill the reconstructive demands and has an edge over the two-stage countermeasure. The authors adopted tripartite SCIAP flaps to reconstruct distal complex thumb or toe injuries in eight cases (six thumbs and two halluces), all of which were reattached with vascularized fascia lata-iliac crest conjunctions using a pull-out technique. All SCIAP flaps survived uneventfully without donor-site complications. The remodeled interphalangeal joints regained nearly normal radiologic manifestation. The chimeric SCIAP flap may be a promising technique for distal complex extensor tendon injury; providing vascularized skin paddle and fascia lata-iliac crest graft, it also qualifies for the all-in-one-stage reconstruction concept. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.

The bone mesenchymal stem cell-derived exosomal miR-146a-5p promotes diabetic wound healing in mice via macrophage M1/M2 polarization.

A diabetic wound is a refractory disease that afflicts patients globally. MicroRNA-146a-5p (miR-146a-5p) is reported to represent a potential therapeutic target for diabetic wounds. However, microRNA easily degrades in the wound microenvironment. This study extracted bone marrow mesenchymal stem cell (BMSC)-derived exosomes (EXO). Electroporation technology was used to load miR-146a-5p into EXO (labeled as EXO-miR-146a). The endothelial cells (human umbilical vein endothelial cells [HUVECs]) and macrophages were cocultured in transwell chambers in the presence of high glucose. Cell proliferation, migration, and angiogenesis were measured with cell counting kit 8, scratch, and tube forming assays, respectively. Flow cytometry was introduced to validate the biomarker of macrophages and BMSCs. The expression level of macrophage polarization-related proteins and tumor necrosis factor receptor-associated factor 6 (TRAF6) was assessed with western blotting analysis. The full-thickness skin wound model was developed to verify the in vitro results. EXO-miR-146a promoted the proliferation, migration, and angiogenesis of HUVECs in the hyperglycemic state by suppressing the TRAF6 expression in vitro. Additionally, EXO-miR-146a treatment facilitated M2 but inhibited M1 macrophage polarization. Furthermore, EXO-miR-146a enhances reepithelialization, angiogenesis, and M2 macrophage polarization, thereby accelerating diabetic wound healing in vivo. The EXO-miR-146a facilitated M2 macrophage polarization, proliferation, migration, and angiogenesis of HUVECs through TRAF6, thereby ameliorating intractable diabetic wound healing. These results established the basis for using EXO to deliver drugs and revealed mediators for diabetic wound treatment.

Receptor Plants Alleviated Allelopathic Stress from Invasive Chenopodium ambrosioides L. by Upregulating the Production and Autophagy of Their Root Border Cells.

Chenopodium ambrosioides L. is an invasive plant native to the Neotropics that has seriously threatened the ecological security of China, and allelopathy is one of the mechanisms underlying its successful invasion. Maize (Zea mays L.) and soybean (Glycine max (L.) Merr.), as the main food crops, are usually affected by C. ambrosioides in their planting areas. The purpose of this study was to investigate the ultrastructure, autophagy, and release-related gene expression of receptor plant root border cells (RBCs) after exposure to volatile oil from C. ambrosioides and its main component α-terpene, which were studied using maize and soybean as receptor plants. The volatiles inhibited root growth and promoted a brief increase in the number of RBCs. As the volatile concentration increased, the organelles in RBCs were gradually destroyed, and intracellular autophagosomes were produced and continuously increased in number. Transcriptomic analysis revealed that genes involved in the synthesis of the plasma membrane and cell wall components in receptor root cells were significantly up-regulated, particularly those related to cell wall polysaccharide synthesis. Meanwhile, polygalacturonase and pectin methylesterases (PME) exhibited up-regulated expression, and PME activity also increased. The contribution of α-terpene to this allelopathic effect of C. ambrosioides volatile oil exceeded 70%. Based on these results, receptor plant root tips may increase the synthesis of cell wall substances while degrading the intercellular layer, accelerating the generation and release of RBCs. Meanwhile, their cells survived through autophagy of RBCs, indicating the key role of RBCs in alleviating allelopathic stress from C. ambrosioides volatiles.

The competitive strategies of poisonous weeds Elsholtzia densa Benth. on the Qinghai Tibet Plateau: Allelopathy and improving soil environment.

Introduction: The competitive strategies of plants play a crucial role in their growth. Allelopathy is one of the weapons that plants use to improve their competitive advantage. Methods: In order to explore the competitive strategy of a poisonous weed Elsholtzia densa Benth. (E. densa) on the Qinghai-Tibet Plateau (QTP), the effects of decomposing substances of E. densa on growth, root border cells (RBCs) characteristics of highland crop highland barley (Hordeum vulgare L.), and soil environment were determined. Results: The decomposing allelopathic effect of E. densa on the germination and seedling growth of highland barley mainly occurred in the early stage of decomposing. The allelopathic effects were mainly on seed germination and root growth of highland barley. After treatment with its decomposing solution, the RBC's mucilage layer of highland barley thickened, and the RBC's activity decreased or even apoptosis compared with the control. However, only the above-ground part of the treatment group showed a significant difference. The effects of E. densa decomposed substances on the soil environment were evaluated from soil physicochemical properties and bacterial community. The results showed that soil bacteria varied greatly in the early stage of decomposion under different concentrations of E. densa. In addition, E. densa decomposing substances increased the soil nutrient content, extracellular enzyme activities, and bacterial community diversity. In the process of decomposition, the bacterial community structure changed constantly, but Actinobacteriota was always the dominant phylum. Discussion: These results indicated that E. densa might adopt the following two strategies to help it gain an advantage in the competition: 1. Release allelochemicals that interfere with the defense function of surrounding plants and directly inhibit the growth and development of surrounding plants. 2. By changing the physical and chemical properties of soil and extracellular enzyme activity, residual plant decomposition can stimulate soil microbial activity, improve soil nutrition status, and create a more suitable soil environment for growth.

Protein kinase D1 promotes the survival of random-pattern skin flaps in rats.

BACKGROUND: In reconstructive surgery, random skin flaps are commonly used tools to cover skin defects, however, their applicability and size are limited by post-operative complications such as marginal ischemia-reperfusion injury and flap necrosis. Protein kinase D1 (PKD1), a calcium/calmodulin-dependent serine/threonine kinase, is known to induce angiogenesis and has been shown to mitigate ischemia in cardiovascular diseases. However, the role of PKD1 has not been investigated in skin flaps. METHOD: Seventy-five male Sprague-Dawley rats with skin flaps were randomly divided into three groups: control, PKD1, and CID755673. Seven days following surgery, we assessed the general view and survival rate of the flap using histological analysis. Laser Doppler and lead oxide/gelatin angiography were used to evaluate microcirculation blood flow. Histopathological changes, neovascularization and microvascular density (MVD). were examined and calculated using microscopy after H&E staining. Protein expression levels were determined using immunoblotting and immunohistochemistry techniques. RESULT: PKD1 significantly improved flap survival by upregulating angiogenic factors VEGF and cadherin5 and increasing antioxidant enzymes SOD, eNOS, and HO1, as well as reducing caspase 3, cytochrome c, and Bax expression, and attenuating IL-1ß, IL-6, and TNF-α. In the PKD1 group, PKD1 increased neovascularization, and blood flow and flap survival areas were larger as compared to the control and CID755673 groups. CONCLUSION: These findings show that PKD1 accelerates angiogenesis, reduces oxidative stress, and impedes apoptosis and inflammation, thus resulting in improved flap survival. Our observations indicated that PKD1 could be a therapeutic target for flap failure treatment.

Climate-induced habitat suitability changes intensify fishing impacts on the life history of large yellow croaker (Larimichthys crocea).

Intense fishing pressure and climate change are major threats to the fish population and coastal fisheries. Larimichthys crocea (large yellow croaker) is a long-lived fish, which performs seasonal migrations from its spawning and nursery grounds along the coast of the East China Sea (ECS) to overwintering grounds offshore. This study used length-based analysis and habitat suitability index (HSI) model to evaluate the current life-history parameters and overwintering habitat suitability of L. crocea, respectively. We compared recent (2019) and historical (1971-1982) life-history parameters and overwintering HSI to analyze the fishing pressure and climate change effects on the overall population and overwintering phase of L. crocea. The length-based analysis indicated serious overfishing of L. crocea, characterized by reduced catch, size truncation, constrained distribution, and advanced maturation causing a recruitment bottleneck. The overwintering HSI modeling results indicated that climate change has led to decreased sea surface temperature during L. crocea overwintering phase over the last half-century, which in turn led to area decrease and an offshore-oriented shifting of optimal overwintering habitat of L. crocea. The fishing-caused size truncation may have constrained the migratory ability, and distribution of L. crocea subsequently led to the mismatch of the optimal overwintering habitat against climate change background, namely habitat bottleneck. Hence, while heavy fishing was the major cause of L. crocea collapse, climate-induced overwintering habitat suitability may have intensified the fishery collapse of L. crocea population. It is important for management to consider both overfishing and climate change issues when developing stock enhancement activities and policy regulations, particularly for migratory long-lived fish that share a similar life history to L. crocea. Combined with China's current restocking and stock enhancement initiatives, we propose recommendations for the future restocking of L. crocea in China.

Risk Factors Associated With Misdiagnosis of Digital Glomus Tumor: A Retrospective Cohort Study.

OBJECTIVE: Glomus tumors are benign with unique triad of symptoms; however, the delayed diagnosis of these tumors is common. We investigated the possible risk factors for the misdiagnosis of digital glomus tumors, with an aim to treat these patients on time. METHODS: We conducted a retrospective cohort study of 104 patients with digital glomus tumors from October 2009 to February 2021. Data pertaining to sex, age, tumor locations, symptoms, imaging modalities, and clinical departments visited by the patients were extracted and analyzed through logistic regression. RESULTS: The duration of delayed diagnosis ranged from 3 months to 40 years (mean, 5.5 ± 6.5 years). The total misdiagnosis and recurrence rate are 34.6% and 3.8%, respectively. On the multivariate logistic regression, the misdiagnosis of digital glomus tumor was significantly associated with the clinical departments visited by the patients ( P < 0.001). The risk of misdiagnosis of nonhand surgery department visit is 179.741-fold higher than that of hand surgery department visit. CONCLUSIONS: The misdiagnosis rate of digital glomus tumor was closely related to the clinical departments visited by the patients. Hand surgeons are the first choice for the treatment of the tumor.

Melatonin promotes peripheral nerve repair through Parkin-mediated mitophagy.

Schwann cells (SCs) are the major glial cells in peripheral nervous system. They unsheathe and myelinate axons and play an essential role in peripheral nerve regeneration. Several studies report that Parkin-mediated mitophagy is associated with various diseases. Melatonin promotes proliferation of central glial cells. Little is known about the effect of melatonin and Parkin-mediated mitophagy on peripheral nerve repair. In this study, using a rat model of a peripheral nerve injury (PNI) and in vitro model established by RSC96 cells treated with tert-butyl hydroperoxide (TBHP), we found that Parkin-mediated mitophagy can effectively reduce the production of mitochondrial reactive oxygen species (ROS), maintain the balance of mitochondrial membrane potential, maintain autophagic flux, and inhibit mitochondrial apoptosis. At the same time, we found that the increase of Parkin under stress is a manifestation of the RSC96 cells' resistance to oxidative stress to maintain RSC96 cells' balance. In our experiment, melatonin is similar to a Parkin agonist, up-regulating the expression of Parkin, enhancing all the positive results of Parkin in a stress state, such as inhibiting active oxygen production, maintaining autophagic flux, and inhibiting mitochondrial apoptosis. In addition, we design in vivo experiments to verify in In vitro experiments. In in vivo, melatonin promotes the expression of Parkin, maintains autophagic flux, inhibits apoptosis, promotes myelin regeneration, reduces the regeneration of collagen fibers around damaged tissues, and promotes peripheral nerve repair. When adenovirus was used to down-regulate the expression of Parkin, we found that all the positive effects of melatonin were attenuated. Collectively, these findings indicate that melatonin upregulates Parkin-mediated mitophagy and promotes peripheral nerve repair. The results provide a basis for development of effective drugs for PNI treatment.

Exploring the Correlation Between the Regulation of Macrophages by Regulatory T Cells and Peripheral Neuropathic Pain.

OBJECTIVE: Intractable pain after peripheral nerve injury has become a major concern in the field of pain. Current evidence shows that routine medications or surgical treatment is associated with inconsistent results and different curative effects. Stable and effective treatment methods in clinical practice are also lacking. To date, there is no consensus on the pathophysiological mechanisms of pain. The present study investigates the potential regulatory role of regulatory T cells in the differentiation of macrophages on dorsal root ganglion (DRG) and explores the mechanism of nociceptive signals in the signal transfer station. The findings are expected to guide the prevention of various types of peripheral neuropathic pain. METHODS: Thirty-six male Sprague Dawley (SD) rats and 18 male Nude rats, of equal weight (250-300g), were used in this study. The rats were divided into 3 groups: SD rat sciatic nerve transection group (SNT group, n = 18), SD rat nerve transection experimental group (SNT/RAPA group, n = 18) and Nude rat nerve transection experimental group (SNT/NUDE group, n = 18). The behavior related to neuropathic pain of animals were comprehensively evaluated in all groups. Furthermore, we analyzed the degree of neuroma development, histology, gene, and protein expression, and compared their correlation with the ultrastructural changes of M1/M2 type differentiation of macrophages in DRG. RESULTS: Sciatic nerve transection (SNT), induced the aggregation of several types of macrophages in lumbar DRG of SD rats leading to a higher ratio of M1/M2. Following the inhibition of the M1 type polarization of macrophages, axon outgrowth increased significantly. A significantly lower average autotomy score was reported in the SNT/NUDE group (*p < 0.05) and the SNT/RAPA group (@ p < 0.05) as compared to that of the SNT group. The SNT/NUDE group showed no noticeable neuroma formation 30 days after the nerve transection. However, bulbous neuromas were observed in the nerve stumps of both the SNT control and SNT/RAPA groups. Immunofluorescence staining revealed a significant decrease in the proportion of M1/M2 macrophages in lumbar DRG of the SNT/NUDE group (** p < 0.001) and the SNT/RAPA group (@ p < 0.05) compared to the SNT group. The expression of pain-related proteins was also decreased (@ p < 0.05, *p < 0.05,** p < 0.001). Also, the expression of alpha-smooth muscle actin (α-SMA), neurofilament 200 (NF-200), and nerve growth factor low-affinity receptor p75 were significantly down-regulated in the nerve tissue (@ p < 0.05, @@ p < 0.001, ** p < 0.001). CONCLUSION: M1/M2 type differentiation of macrophages on DRG plays a significant role in the formation of traumatic painful neuroma after neurotomy. In combination with our previous study, the results of this study suggest that regulatory T cells reduce the ratio of M1/M2 macrophages and alleviate the pain of neuroma by regulating the polarization direction of macrophages on neuroma. These findings provide key insights into developing new strategies to manage painful neuroma.

A new insight on peripheral nerve repair: the technique of internal nerve splinting.

OBJECTIVE: Neuropathic pain produced by symptomatic neuromas is an important problem after peripheral nerve injury (PNI). End-to-end anastomosis of the nerve stump for PNI is well established but cannot efficiently prevent neuroma-in-continuity formation. METHODS: Sciatic nerve injury was used in the experimental model. Seventy-two rats were randomly divided into four groups: rats with nerve anastomosis sites supported with silicone tubes represented the internal nerve splinting (INS) group (n = 18); rats with end-to-end nerve anastomosis represented control group 1 (CON1) (n = 18); rats with INS and the nerve anastomosis site represented control group 2 (CON2) (n = 18); and rats that underwent the same surgical procedures for skin and muscle operations but without sciatic nerve injury represented the normal group (n = 18). RESULTS: Gross evaluations of the nerve anastomosis sites, gastrocnemius muscle atrophy, axonal regeneration and remyelination, neuropathic pain, and scar hyperplasia of the neuromas were performed, as well as motor function evaluations. Axonal regeneration, remyelination, and gastrocnemius muscle atrophy were similar between the INS group and CON1 (p > 0.05). However, neuropathic pain and scar hyperplasia-as evaluated according to the expression of anti-sigma-1 receptor antibody and anti-α-smooth muscle actin, respectively-and the weight ratios of the neuromas were reduced in the INS group compared with those of CON1 and CON2 (p < 0.05). CONCLUSIONS: Application of INS in nerve repair effectively prevented traumatic neuroma-in-continuity formation and inhibited neuropathic pain without influencing nerve regeneration in rats.

Augmenting Peripheral Nerve Regeneration with Adipose-Derived Stem Cells.

Peripheral nerve injuries (PNIs) are common and debilitating, cause significant health care costs for society, and rely predominately on autografts, which necessitate grafting a nerve section non-locally to repair the nerve injury. One possible approach to improving treatment is bolstering endogenous regenerative mechanisms or bioengineering new nervous tissue in the peripheral nervous system. In this review, we discuss critical-sized nerve gaps and nerve regeneration in rats, and summarize the roles of adipose-derived stem cells (ADSCs) in the treatment of PNIs. Several regenerative treatment modalities for PNI are described: ADSCs differentiating into Schwann cells (SCs), ADSCs secreting growth factors to promote peripheral nerve growth, ADSCs promoting myelination growth, and ADSCs treatments with scaffolds. ADSCs' roles in regenerative treatment and features are compared to mesenchymal stem cells, and the administration routes, cell dosages, and cell fates are discussed. ADSCs secrete neurotrophic factors and exosomes and can differentiate into Schwann cell-like cells (SCLCs) that share features with naturally occurring SCs, including the ability to promote nerve regeneration in the PNS. Future clinical applications are also discussed.

Macrophage Activation in the Dorsal Root Ganglion in Rats Developing Autotomy after Peripheral Nerve Injury.

Autotomy, self-mutilation of a denervated limb, is common in animals after peripheral nerve injury (PNI) and is a reliable proxy for neuropathic pain in humans. Understanding the occurrence and treatment of autotomy remains challenging. The objective of this study was to investigate the occurrence of autotomy in nude and Wistar rats and evaluate the differences in macrophage activation and fiber sensitization contributing to the understanding of autotomy behavior. Autotomy in nude and Wistar rats was observed and evaluated 6 and 12 weeks after sciatic nerve repair surgery. The numbers of macrophages and the types of neurons in the dorsal root ganglion (DRG) between the two groups were compared by immunofluorescence studies. Immunostaining of T cells in the DRG was also assessed. Nude rats engaged in autotomy with less frequency than Wistar rats. Autotomy symptoms were also relatively less severe in nude rats. Immunofluorescence studies revealed increased macrophage accumulation and activation in the DRG of Wistar rats. The percentage of NF200+ neurons was higher at 6 and 12 weeks in Wistar rats compared to nude rats, but the percentage of CGRP+ neurons did not differ between two groups. Additionally, macrophages were concentrated around NF200-labeled A fibers. At 6 and 12 weeks following PNI, CD4+ T cells were not found in the DRG of the two groups. The accumulation and activation of macrophages in the DRG may account for the increased frequency and severity of autotomy in Wistar rats. Our results also suggest that A fiber neurons in the DRG play an important role in autotomy.

Aligned nanofiber nerve conduits inhibit alpha smooth muscle actin expression and collagen proliferation by suppressing TGF-ß1/SMAD signaling in traumatic neuromas.

Transforming growth factor-beta 1 (TGF-ß1) is a powerful activator of connective tissue synthesis that is strongly associated with the pathophysiology of traumatic neuroma. Previous studies have demonstrated that aligned nanofiber conduits made from silk fibroin and poly (L-lactic acid-co-ε-caprolactone; PLCL) could prevent traumatic neuromas. In the present study, the possible mechanisms of conduits in treating traumatic neuromas were investigated to provide theoretical basis for procedures. Aligned nanofiber conduits were used for nerve capping. Sciatic nerves of Sprague-Dawley rats were used to create an animal model. The present study contains two parts, each including four experimental groups. SB-431542/SRI-011381 hydrochloride was used to suppress/enhance TGF-ß1/SMAD signaling. Part I discussed the connections between traumatic neuroma and the proliferation of alpha smooth muscle actin (α-SMA) and collagen; it also investigated the therapeutic effect of conduits. Part II hypothesized that conduits suppressed TGF-ß1/SMAD signaling. Histological characteristics, quantitative analysis of α-SMA, collagens and signaling-related parameters were assessed and compared among groups one month postoperatively. Results from Part I demonstrated that aligned nanofiber conduits suppressed the expression of α-SMA and collagens; and results from Part II revealed the downregulation of pathway-related proteins, suggesting that the suppression was mediated by TGF-ß1/SMAD signaling. Aligned nanofiber conduits may be effective nerve capping biomaterials. One of the mechanisms involves suppressing TGF-ß1/SMAD signaling. Novel treatments using aligned nanofiber conduits could be developed to manage traumatic neuromas.

Identification of sensory and motor nerve fascicles by immunofluorescence staining after peripheral nerve injury.

BACKGROUND: Inappropriate matching of motor and sensory fibers after nerve repair or nerve grafting can lead to failure of nerve recovery. Identification of motor and sensory fibers is important for the development of new approaches that facilitate neural regeneration and the next generation of nerve signal-controlled neuro-prosthetic limbs with sensory feedback technology. Only a few methods have been reported to differentiate sensory and motor nerve fascicles, and the reliability of these techniques is unknown. Immunofluorescence staining is one of the most commonly used methods to distinguish sensory and motor nerve fibers, however, its accuracy remains unknown. METHODS: In this study, we aim to determine the efficacy of popular immunofluorescence markers for motor and sensory nerve fibers. We harvested the facial (primarily motor fascicles) and sural (primarily sensory fascicles) nerves in rats, and examined the immunofluorescent staining expressions of motor markers (choline acetyltransferase (ChAT), tyrosine kinase (TrkA)), and sensory markers [neurofilament protein 200 kDa (NF-200), calcitonin gene-related peptide (CGRP) and Transient receptor potential vanillic acid subtype 1 (TRPV1)]. Three methods, including the average area percentage, the mean gray value, and the axon count, were used to quantify the positive expression of nerve markers in the immunofluorescence images. RESULTS: Our results suggest the mean gray value method is the most reliable method. The mean gray value of immunofluorescence in ChAT (63.0 ± 0.76%) and TRKA (47.6 ± 0.43%) on the motor fascicles was significantly higher than that on the sensory fascicles (ChAT: 49.2 ± 0.72%, P < 0.001; and TRKA: 29.1 ± 0.85%, P < 0.001). Additionally, the mean gray values of TRPV1 (51.5 ± 0.83%), NF-200 (61.5 ± 0.62%) and CGRP (37.7 ± 1.22%) on the motor fascicles were significantly lower than that on the sensory fascicles respectively (71.9 ± 2.32%, 69.3 ± 0.46%, and 54.3 ± 1.04%) (P < 0.001). The most accurate cutpoint occurred using CHAT/CRCP ratio, where a value of 0.855 had 100% sensitivity and 100% specificity to identify motor and sensory nerve with an area under the ROC curve of 1.000 (P < 0.001). CONCLUSIONS: A combination of ChAT and CGRP is suggested to distinguish motor and sensory nerve fibers.

Therapeutic effects of peripherally administrated neural crest stem cells on pain and spinal cord changes after sciatic nerve transection.

BACKGROUND: Severe peripheral nerve injury significantly affects patients' quality of life and induces neuropathic pain. Neural crest stem cells (NCSCs) exhibit several attractive characteristics for cell-based therapies following peripheral nerve injury. Here, we investigate the therapeutic effect of NCSC therapy and associated changes in the spinal cord in a sciatic nerve transection (SNT) model. METHODS: Complex sciatic nerve gap injuries in rats were repaired with cell-free and cell-laden nerve scaffolds for 12 weeks (scaffold and NCSC groups, respectively). Catwalk gait analysis was used to assess the motor function recovery. The mechanical withdrawal threshold and thermal withdrawal latency were used to assess the development of neuropathic pain. Activation of glial cells was examined by immunofluorescence analyses. Spinal levels of extracellular signal-regulated kinase (ERK), NF-κB P65, brain-derived neurotrophic factor (BDNF), growth-associated protein (GAP)-43, calcitonin gene-related peptide (CGRP), and inflammation factors were calculated by western blot analysis. RESULTS: Catwalk gait analysis showed that animals in the NCSC group exhibited a higher stand index and Max intensity At (%) relative to those that received the cell-free scaffold (scaffold group) (p < 0.05). The mechanical and thermal allodynia in the medial-plantar surface of the ipsilateral hind paw were significantly relieved in the NCSC group. Sunitinib (SNT)-induced upregulation of glial fibrillary acidic protein (GFAP) (astrocyte) and ionized calcium-binding adaptor molecule 1 (Iba-1) (microglia) in the ipsilateral L4-5 dorsal and ventral horn relative to the contralateral side. Immunofluorescence analyses revealed decreased astrocyte and microglia activation. Activation of ERK and NF-κB signals and expression of transient receptor potential vanilloid 1 (TRPV1) expression were downregulated. CONCLUSION: NCSC-laden nerve scaffolds mitigated SNT-induced neuropathic pain and improved motor function recovery after sciatic nerve repair. NCSCs also protected the spinal cord from SNT-induced glial activation and central sensitization.

The feasibility and survival mechanism of a large free flap supported by a novel hybrid perfusion mode.

OBJECTIVES: In this study, we presented a novel hybrid perfusion mode in an attempt to provide a new strategy to improve the survival of an extended large flap and discuss its possible mechanisms. MATERIALS AND METHODS: A 14 × 10 cm flap was designed on the rabbit abdomen. Ninety-six rabbits were randomly divided into three groups based on the flap perfusion mode: control group I (CON 1, physiological perfusion mode with bilateral deep inferior epigastric vascular pedicles intact), control group II (CON 2, physiological perfusion mode with single deep inferior epigastric vascular pedicle intact), hybrid nourished group (physiological perfusion as in CON 2 combined with arterialized venous nonphysiological perfusion mode, referred to as a hybrid perfusion mode). Flap survival, status of vascular perfusion, microvasculature, histopathology, expression of CD34, eNOs, VEGF and metabolic status of the flaps by LC-MS were assessed in each group. RESULTS: The results of "hybrid nourished" flaps were similar to the traditional flaps in terms of flap survival, level of vascular perfusion and microvasculature except the status of metabolites. CONCLUSIONS: The feasibility of this novel hybrid perfusion mode will greatly extend the indications of flap transfer and efficiently improve the survival reliability of large flaps. In a sense, this mode will be an ideological emancipation for the field of flap surgery.

Risk Factors Associated With Postoperative Recurrence in Patients With Tenosynovial Giant Cell Tumor of the Hand: A Retrospective Cohort Study.

Identification of risk factors for recurrence of tenosynovial giant cell tumors of the hand is crucial to provide adequate preoperative counseling and tailor surgical treatment. However, the risk factors are still controversial, which are the subject of this research.Recently, we conducted a retrospective cohort study of 135 consecutive patients with giant cell tumors of the tendon sheath of the hand from January 2010 to July 2016. All patients underwent surgical excision, received necessary imaging examinations, and had routine follow-up and thus were identified as those who had recurrence by confirmation of reoperation, and the duration ranged from 24 to 103 months (mean, 53.5 ± 21.4 months). There were 14 local recurrences (10.4%) within 6 to 24 months, respectively, after surgery. Data pertaining to sex, age, tumor sites, tumor size, tumor number, course of disease, bone erosion, tumor growth patterns, anesthesia mode, and the surgeon's experience were all extracted, and Cox regression models were used to estimate recurrence rate with adjustment for potential confounders.According to the Cox regression analysis, the recurrence rate after surgery was significantly higher in patients with a diffused form than in those with a localized one (P = 0.001); in addition, patients with 2 or more tumors had a much higher postoperative recurrence rate than did those with only 1 tumor (P = 0.023).This study suggested that the recurrence rate of tenosynovial giant cell tumors of the hand was closely related to the tumor number and tumor growth patterns.

The management of degloving injuries of the limb with full thickness skin grafting using vacuum sealing drainage or traditional compression dressing: A comparative cohort study.

BACKGROUND: Degloving injuries of the limb are severe and frequently underrated. Few researches are available comparing the results of reattachment of the degloved skin grafts with the vacuum sealing drainage technique and the traditional compression dressing method. In this study, we aimed to compare the treatment outcomes of these two approaches. METHODS: Eighty-three consecutive patients were treated for degloving injury of the limb. Based on the treatment approach, the patients were divided into vacuum sealing drainage group (VSD group, n = 55) and traditional compression dressing group (TCD group, n = 28). After reattachment, the degloved skin was secured with stitches and compressed with VSD or TCD. The outcomes were mainly assessed based on the percentage of skin graft take. RESULT: In VSD group, there were excellent results in 18, fair in 9 and poor in 28, respectively; In TCD group, there were excellent results in 11, fair in 10 and poor in 7, respectively. Statistically, no significant difference was found between two groups in terms of the category of excellent results. However, significant higher incidence of poor results with necrotic areas exceeding 50% was observed in the VSD group than that in the TCD group. In addition, although the duration of hospitalization in the VSD group was shorter than that in the TCD group, the medical supply costs and total costs were much higher than that of the TCD group. CONCLUSION: VSD and TCD are equally effective in the management of degloving injuries of the limb; however, VSD technique may potentially have a higher risk of poor results with increased hospital charges. The traditional approach still has its merits in clinical practice, especially in rural hospitals when VSD is not available or unaffordable.

An aligned nanofiber nerve conduit that inhibits painful traumatic neuroma formation through regulation of the RhoA/ROCK signaling pathway.

OBJECTIVE: Traumatic neuromas represent a prevalent source of neuropathic pain. As of yet, there has been no single treatment method that can guarantee permanent relief of symptoms. Although nerve-capping techniques have shown promise, their exact mechanisms remain elusive. The authors' aim was to examine the role of the RhoA/ROCK signaling pathway in the prevention of neuroma formation after neurectomy utilizing a nerve-capping technique. METHODS: An aligned nanofiber tube was fabricated to cap the sciatic nerve in Sprague Dawley rats. The rats (n = 60) were randomly divided into the aligned SF/P (LLA-CL) capping group (capping group, n = 20), the capping and Y-27632 (ROCK pathway inhibitor) intervention group (intervention group, n = 20), and the no-capping group (control group, n = 20). The authors undertook a comprehensive assessment of the capping group, examining the animals' behavior, the extent of neuroma development, histology, gene and protein expression, and ultrastructural changes associated with the RhoA/ROCK signaling pathway. These findings were compared with those in the intervention and control groups. RESULTS: The inciting injury resulted in the expression of the RhoA/ROCK signaling pathway, as well as its further upregulation in peripheral neurons. Axon outgrowth was significantly increased when RhoA/ROCK signaling pathway was suppressed. The average autotomy score in the capping group was observed to be much lower than that of the intervention and control groups. At 30 days postneurectomy, the capping group displayed no obvious neuroma formation, while a bulbous neuroma was found in the nerve stumps of both the control and intervention groups. Quantitative real-time polymerase chain reaction and the Western blot analysis demonstrated that the expression of myelin-associated glycoprotein was substantially upregulated in the capping group; in contrast, the expression of NF-200 was significantly downregulated. The expression of myosin light chain was notably lower in the intervention group, but there was no significant difference when compared with the control group (p > 0.05). CONCLUSIONS: The RhoA/ROCK signaling pathway has emerged as a critical player in the process of traumatic neuroma formation after neurectomy. It is possible that the nerve-capping technique could generate a "regenerative brake" based on the regulation of the RhoA/ROCK signaling pathway in this event. These findings may provide concrete evidence that could help develop new strategies for the management of painful neuromas.