Reendothelialization of Acellular Adipose Flaps under Mimetic Physiological Dynamic Conditions.

The extensive soft-tissue defects resulting from trauma and tumors pose a prevalent challenge in clinical practice, characterized by a high incidence rate. Autologous tissue flap transplantation, considered the gold standard for treatment, is associated with various drawbacks, including the sacrifice of donor sources, postoperative complications, and limitations in surgical techniques, thereby impeding its widespread applicability. The emergence of tissue-engineered skin flaps, notably the acellular adipose flap (AAF), offers potential alternative solutions. However, a critical concern confronting large-scale tissue-engineered skin flaps currently revolves around the reendothelialization of internal vascular networks. In our study, we have developed an AAF utilizing perfusion decellularization, demonstrating excellent physical properties. Cytocompatibility experiments have confirmed its cellular safety, and cell adhesion experiments have revealed spatial specificity in facilitating endothelial cells adhesion within the adipose flap scaffold. Using a novel mimetic physiological fluid shear stress setting, endothelial cells were dynamically inoculated and cultured within the acellular vascular network of the pedicled AAF in our research. Histological and gene expression analyses have shown that the mimetic physiological fluid dynamic model significantly enhanced the reendothelialization of the AAF. This innovative platform of acellular adipose biomaterials combined with hydrodynamics may offer valuable insights for the design and manufacturing of 3D vascularized tissue constructs, which can be applied to the repair of extensive soft-tissue defects.

Cytokine status and significant increase of IL-6 and sIL-6R in the aqueous humor of diabetic cataract patients revealed by quantitative multiplexed assays.

OBJECTIVE: This study aimed to investigate inflammatory cytokine expression profiles in the aqueous humor (AH) of diabetic cataract (DC) patients. METHODS: A quantitative multiplexed antibody assay was performed to measure the expression levels of 40 inflammatory cytokines in AH samples from DC and age-related cataract (ARC) patients. Bioinformatics analysis was used to examine the functions of the cytokines. Enzyme-linked immunosorbent assays (ELISAs) and western blots were performed to verify the data. RESULTS: The multiplexed antibody assay revealed that the expression levels of IL-6, sIL-6R, IL-17A, IL-8, MCP-1, TNF-ß, RANTES, TIMP-1, and TIMP-2 were higher in the AH of DC patients compared with ARC patients. However, IL-1ra and IL-1a expression levels were lower in the DC patient AH samples. Pathway analysis indicated that IL-6 and sIL-6R belong to the class I helical cytokine family, which is associated with many biological functions. ELISA and western blot results confirmed that IL-6R and IL-6 expression levels were significantly higher in DC patients compared with ARC patients. CONCLUSIONS: Our results revealed the status of 40 inflammatory cytokines in the AH by quantitative multiplexed assays. Additionally, IL-6 and sIL-6R were expressed markedly higher in DC compared with ARC, which may play critical roles in DC pathophysiology.

Fibulin-7 C-terminal fragment and its active synthetic peptide suppress choroidal and retinal neovascularization.

Wet age-related macular degeneration (AMD) and diabetic retinopathy are the leading causes of blindness through increased angiogenesis. Although VEGF-neutralizing proteins provide benefit, inconsistent responses indicate a need for new therapies. We previously identified the Fibulin-7 C-terminal fragment (Fbln7-C) as an angiogenesis inhibitor in vitro. Here we show that Fbln7-C inhibits neovascularization in vivo, in both a model of wet AMD involving choroidal neovascularization (CNV) and diabetic retinopathy involving oxygen-induced ischemic retinopathy. Furthermore, a short peptide sequence from Fbln7-C is responsible for the anti-angiogenic properties of Fbln7-C. Our work suggests Fbln7-C as a therapeutic candidate for wet AMD and ischemic retinopathy.

Nanofiber-reinforced decellularized amniotic membrane improves limbal stem cell transplantation in a rabbit model of corneal epithelial defect.

Human amniotic membrane (AM) offers unique advantages as a matrix to support the transplantation of limbal stem cells (LSCs) due to its inherent pro-regenerative and anti-inflammatory properties. However, the widespread use of AM in clinical treatments of ocular surface disorders is limited by its weak mechanical strength and fast degradation, and high cost associated with preserving freshly isolated AM. Here we constructed a composite membrane consisting of an electrospun bioabsorbable poly(ε-caprolactone) (PCL) nanofiber mesh to significantly improve the ultimate tensile strength, toughness, and suture retention strength by 4-10-fold in comparison with decellularized AM sheet. The composite membrane showed extended stability and conferred longer-lasting coverage on wounded cornea surface compared with dAM. The composite membrane maintained the pro-regenerative and immunomodulatory properties of dAM, promoted LSC survival, retention, and organization, improved re-epithelialization of the defect area, and reduced inflammation and neovascularization. This study demonstrates the translational potential of our composite membrane for stem cell-based treatment of ocular surface damage. STATEMENT OF SIGNIFICANCE: Human decellularized amniotic membrane (dAM) has been widely shown as a biodegradable and bioactive matrix for regenerative tissue repair. However, the weak mechanical property has limited its widespread use in the clinic. Here we constructed a composite membrane using a layer of electrospun poly(ε-caprolactone) (PCL) nanofiber mesh to reinforce the dAM sheet through covalent interfacial bonding, while retaining the unique bioactivity of dAM. In a rabbit model of limbal stem cell (LSC) deficiency induced by alkaline burn, we demonstrated the superior property of this PCL-dAM composite membrane for repairing damaged cornea through promoting LSC transplantation, improving re-epithelialization, and reducing inflammation and neovascularization. This new composite membrane offers great translational potential in supporting stem cell-based treatment of ocular surface damage.

VEGF/VEGFR2 blockade does not cause retinal atrophy in AMD-relevant models.

Intraocular injections of VEGF-neutralizing proteins provide tremendous benefits in patients with choroidal neovascularization (NV) due to age-related macular degeneration (AMD), but during treatment some patients develop retinal atrophy. Suggesting that VEGF is a survival factor for retinal neurons, a clinical trial group attributed retinal atrophy to VEGF suppression and cautioned against frequent anti-VEGF injections. This recommendation may contribute to poor outcomes in clinical practice from insufficient treatment. Patients with type 3 choroidal NV have particularly high risk of retinal atrophy, an unexplained observation. Herein we show in mouse models that VEGF signaling does not contribute to photoreceptor survival and functioning: (a) neutralization of VEGFR2 strongly suppresses choroidal NV without compromising photoreceptor function or survival; (b) VEGF does not slow loss of photoreceptor function or death in mice with inherited retinal degeneration, and there is no exacerbation by VEGF suppression; and (c) mice with type 3 choroidal NV develop retinal atrophy due to oxidative damage with no contribution from VEGF suppression. Intraocular injections of VEGF-neutralizing proteins, a highly effective treatment in patients with neovascular AMD, should not be withheld or reduced due to concern that they may contribute to long-term visual loss from retinal atrophy.

Prospective randomized comparison of transumbilical two-port laparoscopic and conventional laparoscopic varicocele ligation.

We have established a novel method named transumbilical two-port laparoscopic varicocele ligation (TTLVL) for varicocele, which is still needed to evaluate. In this study, 90 patients with left idiopathic symptomatic varicoceles of grades II-III according to the Dubin grading system were randomly assigned to TTLVL (n = 45) and conventional laparoscopic varicocele ligation (CLVL) (n = 45). The demographic, intraoperative, postoperative, and follow-up data were recorded and compared between the two groups. All the procedures in the two groups were completed successfully with no intraoperative complications and no conversions to open surgery. No significant difference was found in the operative time, resuming ambulation, bowel recovery, postoperative hospital stay, and postoperative resolution of scrotal pain between the two groups (P > 0.05). However, the postoperative mean visual analog pain scale scores for TTLVL group were all less at 24 h, 48 h, 72 h, and 7 days postoperatively compared to CLVL (P = 0.001, 0.010, 0.006, and 0.027, respectively). The mean patient scar assessment questionnaire score in postoperative month 3 was 29.7 for TTLVL group compared with 32.1 for CLVL group (P < 0.001). There was no testicular atrophy observed in both groups during the follow-up period. The study shows that TTLVL is a safe, feasible, and effective minimally invasive surgical alternative to CLVL for the treatment of varicocele. Compared with CLVL, TTLVL may decrease postoperative pain and improve the cosmetic outcomes.

[Intramedullary nailing combined with cannulated screw in treating femoral condyles fractures].

OBJECTIVE: To observe the clinical effects of retrograde intramedullary nailing and cannulated screws in the treatment of femoral condylar fracture. METHODS: From June 2009 to June 2015, 13 patients with femoral condyles fracture were treated by retrograde intramedullary nailing and cannulated screws including 6 males and 7 females with an average age of 46.1 years old ranging from 16 to 76 years old. There were 10 cases of closed fractures, 3 cases of open fraetures. According to AO classification criteriam, 4 cases were type C1, 7 cases were type C2, 2 cases were type C3. Postoperative reduction of fracture and the knee joint function recovery were observed. RESULTS: All patients were followed up for 12 to 36 months with a mean of 24 months. X-ray examination showed that the union time of fracture was 18 to 24 weeks, 21 weeks on average. There were no cases of loosening, breakage of internal fixators and re-fracture. Hospital for Special Surgery(HSS) knee score was 90.07±4.99 at 1 year after the operation. CONCLUSIONS: The clinical efficacy for retrograde intramedullary nailing and cannulated screw for the treatment of femoral condyles fracture was excellent. It can improve the anatomical reattachment rate and reduce the complications and promote the knee functional recovery.

PPARα Agonist Fenofibrate Ameliorates Learning and Memory Deficits in Rats Following Global Cerebral Ischemia.

Increasing evidence demonstrates that local inflammation contributes to neuronal death following cerebral ischemia. Peroxisome proliferator-activated receptor α (PPARα) activation has been reported to exhibit many pharmacological effects including anti-inflammatory functions. The aim of this study was to investigate the neuroprotective effects of PPARα agonist fenofibrate on the behavioral dysfunction induced by global cerebral ischemia/reperfusion (GCI/R) injury in rats. The present study showed that fenofibrate treatment significantly reduced hippocampal neuronal death, and improved memory impairment and hippocampal neurogenesis after GCI/R. Fenofibrate administration also inhibited GCI/R-induced over-activation of microglia but not astrocytes and prevented up-regulations of pro-inflammatory mediators in hippocampus. Further study demonstrated that treatment with fenofibrate suppressed GCI/R-induced activations of P65 NF-κB and P38 MAPK. Our data suggest that the PPARα agonist fenofibrate can exert functional recovery of memory deficits and neuroprotective effect against GCI/R in rats via triggering of neurogenesis and anti-inflammatory effect mediated by inhibiting activation of P65 NF-κB and P38 MAPK in the hippocampus, which can contribute to improvement in neurological deficits.

Valproic acid alleviates memory deficits and attenuates amyloid-ß deposition in transgenic mouse model of Alzheimer's disease.

In the brains of patients with Alzheimer's disease (AD) and transgenic AD mouse models, astrocytes and microglia activated by amyloid-ß (Aß) contribute to the inflammatory process that develops around injury in the brain. Valproic acid (VPA) has been shown to have anti-inflammatory function. The present study intended to explore the therapeutic effect of VPA on the neuropathology and memory deficits in APPswe/PS1ΔE9 (APP/PS1) transgenic mice. Here, we report that VPA-treated APP/PS1 mice markedly improved memory deficits and decreased Aß deposition compared with the vehicle-treated APP/PS1 mice. Moreover, the extensive astrogliosis and microgliosis as well as the increased expression in interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) in the hippocampus and cortex of APP/PS1 transgenic mice were significantly reduced following administration of VPA, which attenuated neuronal degeneration. Concomitantly, VPA alleviated the levels of p65 NF-κB phosphorylation and enhanced the levels of acetyl-H3, Bcl-2, and phospho-glycogen synthase kinase (GSK)-3ß that occurred in the hippocampus of APP/PS1 transgenic mice. These results demonstrate that VPA could significantly ameliorate spatial memory impairment and Aß deposition at least in part via the inhibition of inflammation, suggesting that administration of VPA could provide a therapeutic approach for AD.

Overexpression of FoxM1 is associated with tumor progression in patients with clear cell renal cell carcinoma.

BACKGROUND: Fork head box M1 (FoxM1) is a proliferation-associated transcription factor essential for cell cycle progression. Numerous studies have documented that FoxM1 has multiple functions in tumorigenesis and its elevated levels are frequently associated with cancer progression. The present study was conducted to investigate the expression of FoxM1 and its prognostic significance in clear cell renal cell carcinoma (ccRCC). Meanwhile, the function of FoxM1 in human ccRCC was further investigated in cell culture models. METHODS: Real-time quantitative PCR, western blot and immunohistochemistry were used to explore FoxM1 expression in ccRCC cell lines and primary ccRCC clinical specimens. FoxM1 expression was knocked down by small interfering RNA (siRNA) in Caki-1 and 786-O cells; proliferation, colony formation, cell cycle, migration, invasion, and angiogenesis were assayed. RESULTS: FoxM1 expression was up-regulated in the majority of the ccRCC clinical tissue specimens at both mRNA and protein levels. Clinic pathological analysis showed that FoxM1 expression was significantly correlated with primary tumor stage (P <0.001), lymph node metastasis (P = 0.01), distant metastasis (P = 0.01), TNM stage (P < 0.001) and histological grade (P = 0.003). The Kaplan-Meier survival curves revealed that high FoxM1 expression was associated with poor prognosis in ccRCC patients (P < 0.001). FoxM1 expression was an independent prognostic marker of overall ccRCC patient survival in a multivariate analysis (P = 0.008). Experimentally, we found that down-regulation of FoxM1 inhibited cell proliferation and induced cell cycle arrest with reduced expression of cyclin B1, cyclin D1, and Cdk2, and increased expression of p21 and p27. Also, down-regulation of FoxM1 reduced expression and activity of matrix metalloproteinase-2 (MMP-2), MMP-9 and vascular endothelial growth factor (VEGF), resulting in the inhibition of migration, invasion, and angiogenesis. CONCLUSIONS: These results suggest that FoxM1 expression is likely to play important roles in ccRCC development and progression, and that FoxM1 is a prognostic biomarker and a promising therapeutic target for ccRCC.

Vitreous amyloidosis in two large mainland Chinese kindreds resulting from transthyretin variant Lys35Thr and Leu55Arg.

OBJECTIVE: To describe the clinical and pathological findings of two large mainland Chinese kindreds with vitreous amyloidosis and associated transthyretin mutation. METHODS: Twenty individuals from two kindreds with vitreous amyloidosis were ascertained. The transtheretin (TTR) gene of each individual was analyzed, and a clinical examination was obtained on the index patient. RESULTS: Vitreous amyloidosis and radiculopathy were the significant findings in affected individuals. Vitrectomy was performed on the severely affected individuals, with resulting postoperative visual acuity of 20/80 to 20/25. Congo red staining demonstrated amyloid in the vitreous specimen. In Case A, DNA sequencing of exon 2 in the TTR gene revealed a base-pair substitution at codon 35, AAG > ACG (Lys35Thr). In Case B, a missense mutation of leucine-to-arginine substitution was identified at amino acid position 55 in exon 3, CTG > CGG (Leu55Arg). CONCLUSIONS: TTR Lys35Thr and Leu55Arg mutations are associated with vitreous amyloidosis. The phenotype is variable, with vitreous opacities occurring earlier, and sometimes as the sole signs of amyloidotic polyneuropathies (FAPs). Vitrectomy improves vision in some patients with vitreous amyloidosis.

WITHDRAWN: Transplantation of bone mesenchymal stem cells containing the nerve growth factor gene in adult rat brain with Fimbria Fornix lesion.

Bone marrow-derived mesenchymal stem cells (BMSCs) are a promising source for cell-based treatment of brain injury, but the therapy of BMSCs is restricted by low cell survival. We examined whether nerve growth factor (NGF) improve BMSCs viability in the brain with Fimbria-Fornix lesion (FF). After transduction of NGF gene via recombinant retroviral vectors, the rat BMSCs were transformed into the NGF-GFP positive BMSCs, nearly 100% of cells expressed NGF. After transplanted into basal forebrain of rat with FF, the NGF-GFP positive BMSCs expressed the exogenous NGF gene in the host brain, and interesting, the survival number of BMSCs in the NGF group was significant more than that of the void plasmid group. Furthermore, the number of choline acetyltransferase (ChAT) immunoreactive neurons of NGF group was also significant higher than those of the void plasmid group (p<0.05) or the PBS group (p<0.01). Performance in the water maze test was improved in these rats in NGF group. These results indicate that NGF increased BMSCs survival in brain with FF, which results in better improvement of brain function than injected with BMSCs alone.

[Damaging nucleus centromedianus thalami for treatment of cancer pain in experimental and clinical aspects].

BACKGROUND & OBJECTIVE: There is no perfect method to control cancer pain. It is reported that nucleus centromedianus thalami plays a crucial role in the analgesia of central nerve system. The authors conducted this study, based on rat experiments, together with the clinical treatment of more than 90 cases involving various cancer pains, to explore the pain-relieving effects after damaging nucleus centromedianus thalami. METHODS: Ten SD rats, whose nucleus centromedianus thalami were damaged by electrolysis, were chosen, and then measured the pain degree by applying electricity to stimulate the tails of the rats. Meanwhile, another 10 rats, whose nucleus centromedianus thalami were not damaged, were chosen as the control group, among whom the same operation procedure as the above mentioned was carried out. The range of pain scale of the rats was measured by the alteration of the electric intensity. A total of 90 cases of intractable cancer pain were treated, including 36 cases of lung cancer, 21 cases of nasopharyngeal carcinoma, 10 cases of intestinal cancer, 8 cases of cancer of pancreas, 8 cases of osteocarcinoma, 4 cases of carcinoma of kidney, 3 cases of hepatocarcinoma. The brain stereotactic technique was used to damage the nucleus centromedianus thalami with radiofrequency coagulation lesions. The 10-grade method recommended by WHO was used to rank pain degree. RESULTS: Pain scale of rats in the first group rose from 0.152+/-0.034 mA prior to the damage to 0.326+/-0.05 afterwards, with a significant difference (P< 0.001), while the pain scale of the control group dropped from 0.142+/-0.027 mA prior to the operation to 0.138+/-0.035 mA afterwards, with no remarkable difference (P > 0.05). To patients with cancer pain, the average pain grade in this study went above 7 scores, but dropped to 0-3 scores after operation, according to the 10 grade method by WHO. A life-long tracing observation indicted that cancer pain in 24 cases relapsed to varying degrees but below 5 scores, the rest of the patients were analgesic persistently, 3 cases among whom lasted for as long as 2 years. The incidence cases of operational complications were 15 of somnolence, 10 of urinary incontinence, 8 of divagation, and 3 of unilateral oculomotor paralysis. These complications released after symptomatic treatments. CONCLUSION: Nucleus centromedianus thalami damage is an effective way to relieve cancer pain, as well as the complications should be paid attention.