Methods, Mechanisms, and Application Prospects for Enhancing Extracellular Vesicle Uptake.

Extracellular vesicles (EVs) are considered to be a new generation of bioinspired nanoscale drug delivery systems due to their low immunogenicity, natural functionality, and excellent biocompatibility. However, limitations such as low uptake efficiency, insufficient production, and inhomogeneous performance undermine their potential. To address these issues, numerous researchers have put forward various methods and applications for enhancing EV uptake in recent decades. In this review, we introduce various methods for the cellular uptake of EVs and summarize recent advances on the methods and mechanisms for enhancing EV uptake. In addition, we provide further understanding regarding enhancing EV uptake and put forward prospects and challenges for the development of EV-based therapy in the future.

The circ_0002538/miR-138-5p/plasmolipin axis regulates Schwann cell migration and myelination in diabetic peripheral neuropathy.

Circular RNAs (circRNAs) play a vital role in diabetic peripheral neuropathy. However, their expression and function in Schwann cells in individuals with diabetic peripheral neuropathy remain poorly understood. Here, we performed protein profiling and circRNA sequencing of sural nerves in patients with diabetic peripheral neuropathy and controls. Protein profiling revealed 265 differentially expressed proteins in the diabetic peripheral neuropathy group. Gene Ontology indicated that differentially expressed proteins were mainly enriched in myelination and mitochondrial oxidative phosphorylation. A real-time polymerase chain reaction assay performed to validate the circRNA sequencing results yielded 11 differentially expressed circRNAs. circ_0002538 was markedly downregulated in patients with diabetic peripheral neuropathy. Further in vitro experiments showed that overexpression of circ_0002538 promoted the migration of Schwann cells by upregulating plasmolipin (PLLP) expression. Moreover, overexpression of circ_0002538 in the sciatic nerve in a streptozotocin-induced mouse model of diabetic peripheral neuropathy alleviated demyelination and improved sciatic nerve function. The results of a mechanistic experiment showed that circ_0002538 promotes PLLP expression by sponging miR-138-5p, while a lack of circ_0002538 led to a PLLP deficiency that further suppressed Schwann cell migration. These findings suggest that the circ_0002538/miR-138-5p/PLLP axis can promote the migration of Schwann cells in diabetic peripheral neuropathy patients, improving myelin sheath structure and nerve function. Thus, this axis is a potential target for therapeutic treatment of diabetic peripheral neuropathy.

Use of Muscle Feeding Arteries as Recipient Vessels for Soft Tissue Reconstruction in Lower Extremities.

Microsurgical free tissue transfer is still playing an important role in lower extremity reconstruction. Finding a suitable recipient artery for anastomosis is critical in the microsurgical procedure, especially in an extensive wound, or in a complex trauma combined with vascular injury. From April 2014 to March 2018, we retrospectively reviewed patients with traumatic/post-traumatic, oncologic, and electrical wounds in the lower extremity. Those treated with muscle feeding artery as recipient vessels were included. The latissimus dorsi (LD) muscle free flap, anterior lateral thigh (ALT) perforator free flap, and deep inferior epigastric perforator (DIEP) free flap were raised. The muscle feeding arteries to vastus lateral muscle and to medial head of gastrocnemius muscle, concomitant veins, and great saphenous vein were used as recipient vessels. Injuries included in the study were caused by tumour in 2 cases, car accident in 3 cases, crushing in 3 cases, burns in one case, and electrical injury in one case. The wound size varied from 14 cm × 6 cm to 30 cm × 20 cm. LD, ALT, and DIEP free flaps were used in 6, 3, and 4 patients, respectively. The muscle feeding arteries to medial head of gastrocnemius muscle, to sartorius muscle, and to vastus lateral muscle were used as recipient arteries in 4, 5, and one patient, respectively. Concomitant and great saphenous veins were used as recipient veins in 10 and 4 patients, respectively. Using the muscle feeding artery is feasible to avoid injury to the main artery and facilitate dissection and anastomoses, particularly when the wound is located proximal to the mid-third of the lower leg.

A Case of Childhood COVID-19 Infection with Pleural Effusion Complicated by Possible Secondary Mycoplasma Pneumoniae Infection.

We report a case of childhood coronavirus disease 2019 infection with pleural effusion complicated by possible secondary Mycoplasma pneumoniae infection. Fever and pulmonary lesions on computed tomography were the early clinical manifestations, and the patient developed nonproductive cough later. The hydrothorax in this coronavirus disease 2019 case was exudative, showing predominantly mature lymphocytes.

Muscle Fiber Type Changes in Lumbrical Muscles at Early Stages of Chronic Nerve Compression.

Chronic nerve compression (CNC) neuropathy is a common disease in the clinic and provokes paraesthesia, or numbness at early stage. The changes in muscle fiber composition and motor nerve terminal morphology in distal muscles were studied in this study. A well-established CNC model was used to assess the changes in the muscles. Behaviors were measured by von Frey filament test. The myosin heavy chain isoforms and neuromuscular junctions (NMJs) were stained by immunofluorescence to show the muscle fiber types composition and motor nerve terminals morphologic changes in the flexor digitorum longus (FDL) and lumbrical muscle. The fiber cross-sectional areas of different muscle fiber types were measured. The small-fiber degeneration of cutaneous nerve fibers was examined by detecting the protein gene product 9.5 (PGP9.5) with immunofluorescence. At 2nd month after compression, the proportion of type I and type II B fibers was markedly decreased, and that of type II A fibers was increased in the lumbrical muscle. There was no significant change in composition of muscle fiber types in FDL and NMJ morphology of FDL and lumbrical muscles. Intra-epidermal nerve fibre density (IENFD) declined at 2nd month after the compression. Our study reveals the morphological changes of the FDL and lumbrical muscle at an early stage of CNC. These findings may be helpful to understand muscle damage and pathophysiological development of the nerve compression, and provide new evidence for early treatment of CNC.

A novel chronic nerve compression model in the rat.

Current animal models of chronic peripheral nerve compression are mainly silicone tube models. However, the cross section of the rat sciatic nerve is not a perfect circle, and there are differences in the diameter of the sciatic nerve due to individual differences. The use of a silicone tube with a uniform internal diameter may not provide a reliable and consistent model. We have established a chronic sciatic nerve compression model that can induce demyelination of the sciatic nerve and lead to atrophy of skeletal muscle. In 3-week-old pups and adult rats, the sciatic nerve of the right hind limb was exposed, and a piece of surgical latex glove was gently placed under the nerve. N-butyl-cyanoacrylate was then placed over the nerve, and after it had set, another piece of glove latex was placed on top of the target area and allowed to adhere to the first piece to form a sandwich-like complex. Thus, a chronic sciatic nerve compression model was produced. Control pups with latex or N-butyl-cyanoacrylate were also prepared. Functional changes to nerves were assessed using the hot plate test and electromyography. Immunofluorescence and electron microscopy analyses of the nerves were performed to quantify the degree of neuropathological change. Masson staining was conducted to assess the degree of fibrosis in the gastrocnemius and intrinsic paw muscles. The pup group rats subjected to nerve compression displayed thermal hypoesthesia and a gradual decrease in nerve conduction velocity at 2 weeks after surgery. Neuropathological studies demonstrated that the model caused nerve demyelination and axonal irregularities and triggered collagen deposition in the epineurium and perineurium of the affected nerve at 8 weeks after surgery. The degree of fibrosis in the gastrocnemius and intrinsic paw muscles was significantly increased at 20 weeks after surgery. In conclusion, our novel model can reproduce the functional and histological changes of chronic nerve compression injury that occurs in humans and it will be a useful new tool for investigating the mechanisms underlying chronic nerve compression.

Transposition of branches of radial nerve innervating supinator to posterior interosseous nerve for functional reconstruction of finger and thumb extension in 4 patients with middle and lower trunk root avulsion injuries of brachial plexus.

This study aimed to investigate the reconstruction of the thumb and finger extension function in patients with middle and lower trunk root avulsion injuries of the brachial plexus. From April 2010 to January 2015, we enrolled in this study 4 patients diagnosed with middle and lower trunk root avulsion injuries of the brachial plexus via imaging tests, electrophysiological examinations, and clinical confirmation. Muscular branches of the radial nerve, which innervate the supinator in the forearm, were transposed to the posterior interosseous nerve to reconstruct the thumb and finger extension function. Electrophysiological findings and muscle strength of the extensor pollicis longus and extensor digitorum communis, as well as the distance between the thumb tip and index finger tip, were monitored. All patients were followed up for 24 to 30 months, with an average of 27.5 months. Motor unit potentials (MUP) of the extensor digitorum communis appeared at an average of 3.8 months, while MUP of the extensor pollicis longus appeared at an average of 7 months. Compound muscle action potential (CMAP) appeared at an average of 9 months in the extensor digitorum communis, and 12 months in the extensor pollicis longus. Furthermore, the muscle strength of the extensor pollicis longus and extensor digitorum communis both reached grade III at 21 months. Lastly, the average distance between the thumb tip and index finger tip was 8.8 cm at 21 months. In conclusion, for patients with middle and lower trunk injuries of the brachial plexus, transposition of the muscular branches of the radial nerve innervating the supinator to the posterior interosseous nerve for the reconstruction of thumb and finger extension function is practicable and feasible.

Shikonin inhibits TNF-α production through suppressing PKC-NF-κB-dependent decrease of IL-10 in rheumatoid arthritis-like cell model.

Shikonin, a major effective component in the Chinese herbal medicine Lithospermum erythrorhizon Sieb., exhibits an anti-inflammatory property towards rheumatoid arthritis (RA), but the potential mechanism is unclear. Our aim was to investigate the mechanism of shikonin on the lipopolysaccharide (LPS)-induced fibroblast-like synoviocyte (LiFLS) inflammation model. Fibroblast-like synoviocytes (FLSs) were treated with 200 µg/ml of LPS for 24 h to establish the RA-like model, LiFLS. FLSs were pretreated with shikonin (0.1-1 µM) for 30 min in the treatment groups. Quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assays were used to detect mRNA and protein levels of interleukin (IL)-10 and tumor necrosis factor (TNF)-α. Signal proteins involved in IL-10 production were analyzed by Western blotting. Shikonin significantly reversed the inhibitory effects of LPS on IL-10 expression in FLSs by inactivating the PKC-NF-κB pathway. In addition, shikonin inhibited LPS-induced TNF-α expression in FLSs, and this effect was markedly diminished by IL-10-neutralizing antibody. The IL-10-mediated suppression of TNF-α transcription was demonstrated by no response to the protein synthesis inhibitor cyclohexamide and no mRNA decay. Shikonin inhibits LPS-induced TNF-α production in FLSs through suppressing the PKC-NF-κB-dependent decrease in IL-10, and this study also highlights the potential application of shikonin in the treatment of RA.

Transposition of Branches of Radial Nerve Innervating Supinator to Posterior Interosseous Nerve for Functional Reconstruction of Finger and Thumb Extension in 4 Patients with Middle and Lower Trunk Root Avulsion Injuries of Brachial Plexus / 华中科技大学学报（医学）（英德文版）

This study aimed to investigate the reconstruction of the thumb and finger extension function in patients with middle and lower trunk root avulsion injuries of the brachial plexus.From April 2010 to January 2015,we enrolled in this study 4 patients diagnosed with middle and lower trunk root avulsion injuries of the brachial plexus via imaging tests,electrophysiological examinations,and clinical confirmation.Muscular branches of the radial nerve,which innervate the supinator in the forearm,were transposed to the posterior interosseous nerve to reconstruct the thumb and finger extension function.Electrophysiological findings and muscle strength of the extensor pollicis longus and extensor digitorum communis,as well as the distance between the thumb tip and index finger tip,were monitored.All patients were followed up for 24 to 30 months,with an average of 27.5 months.Motor unit potentials (MUP) of the extensor digitorum communis appeared at an average of 3.8 months,while MUP of the extensor pollicis longus appeared at an average of 7 months.Compound muscle action potential (CMAP) appeared at an average of 9 months in the extensor digitorum communis,and 12 months in the extensor pollicis longus.Furthermore,the muscle strength of the extensor pollicis longus and extensor digitorum communis both reached grade Ⅲ at 21 months.Lastly,the average distance between the thumb tip and index finger tip was 8.8 cm at 21 months.In conclusion,for patients with middle and lower trunk injuries of the brachial plexus,transposition of the muscular branches of the radial nerve innervating the supinator to the posterior interosseous nerve for the reconstruction of thumb and finger extension function is practicable and feasible.

Verapamil inhibits scar formation after peripheral nerve repair in vivo.

The calcium channel blocker, verapamil, has been shown to reduce scar formation by inhibiting fibroblast adhesion and proliferation in vitro. It was not clear whether topical application of verapamil after surgical repair of the nerve in vivo could inhibit the formation of excessive scar tissue. In this study, the right sciatic nerve of adult Sprague-Dawley rats was transected and sutured with No. 10-0 suture. The stoma was wrapped with gelfoam soaked with verapamil solution for 4 weeks. Compared with the control group (stoma wrapped with gelfoam soaked with physiological saline), the verapamil application inhibited the secretion of extracellular matrix from fibroblasts in vivo, suppressed type I and III collagen secretion and increased the total number of axons and the number of myelinated axons. These findings suggest that verapamil could reduce the formation of scar tissue and promote axon growth after peripheral nerve repair.

TGF-ß1-induced synthesis of collagen fibers in skeletal muscle-derived stem cells.

The aim of this study was to investigate the mechanism of deposition of extracellular matrix induced by TGF-ß1 in skeletal muscle-derived stem cells (MDSCs). Rat skeletal MDSCs were obtained by using preplate technique, and divided into four groups: group A (control group), group B (treated with TGF-ß1, 10 ng/mL), group C (treated with TGF-ß1 and anti-connective tissue growth factor (CTGF), both in 10 ng/mL), and group D (treated with anti-CTGF, 10 ng/mL). The expression of CTGF, collagen type-I (COL-I) and collagen type-III (COL-III) in MDSCs was examined by using RT-PCR, Western blot and immunofluorescent stain. It was found that one day after TGF-ß1 treatment, the expression of CTGF, COL-I and COL-III was increased dramatically. CTGF expression reached the peak on the day 2, and then decreased rapidly to a level of control group on the day 5. COL-I and COL-III mRNA levels were overexpresed on the day 2 and 3 respectively, while their protein expression levels were up-regulated on the day 2 and reached the peak on the day 7. In group C, anti-CTGF could partly suppress the overexpression of COL-I and COL-II induced by TGF-ß1 one day after adding CTGF antibody. It was concluded that TGF-ß1 could induce MDSCs to express CTGF, and promote the production of COL-I and COL-III. In contrast, CTGF antibody could partially inhibit the effect of TGF-ß1 on the MDSCs by reducing the expression of COL-I and COL-III. Taken together, we demonstrated that TGF-ß1-CTGF signaling played a crucial role in MDSCs synthesizing collagen proteins in vitro, which provided theoretical basis for exploring the methods postponing skeletal muscle fibrosis after nerve injury.

[Combined immunosuppression of FK506 and RS-61443 in rat limb allotransplantation].

OBJECTIVE: To study the immunosuppressive effect of combined therapy with FK506 and RS-61443 in rat limb allotransplantation. METHODS: A total of 101 male SD rats were randomly divided into seven groups and used as recipients, and 101 Wistar rats were used as donors. All SD rats were performed limb allotransplantation without using immunosuppressants in control group. In experimental groups (Groups 1-6), the recipients were immunosuppressed with various dosages of FK506, RS-61443 or FK506 + RS61443, after transplantation for 5 weeks. To evaluate the results, we observed circulation of the transplanted limb, the mean rejection time, the histologic grading of skin rejection of limb grafts and the survival time of limb grafts. RESULTS: The control group showed rejection signs (edema and erythema of the skin) after a mean time of 3.36 +/- 1.15 days, and the mean survival time of the allografts was only 7.00 +/- 0.78 days. In the groups only using FK506 or RS-61443, the survival time were prolonged to varying degrees, but rejection occurred even in the period of using drug. As dosage increased, the rejection could not be prevented and the damage to liver and kidney could be induced. In the group using FK506 in combination with RS-61443, only skin and muscle of limb allografts showed slight rejection sign, function of liver and kidney was not obviously affected, the mean survival time of limb allografts was prolonged to 58.76 +/- 6.81 days. CONCLUSIONS: A combination of FK506 and RS-61443 is a more potent immunosuppressive agent than FK506 oro RS-61443 in preventing the rejection of limb allografts, and it can obviously prolong the survival time of limb allografts.