Amelioration of ligamentum flavum hypertrophy using umbilical cord mesenchymal stromal cell-derived extracellular vesicles.

Ligamentum flavum (LF) hypertrophy (LFH) has been recognised as one of the key contributors to lumbar spinal stenosis. Currently, no effective methods are available to ameliorate this hypertrophy. In this study, human umbilical cord mesenchymal stromal cell-derived extracellular vesicles (hUCMSC-EVs) were introduced for the first time as promising vehicles for drug delivery to treat LFH. The downregulation of miR-146a-5p and miR-221-3p expressions in human LF tissues negatively correlated with increased LF thickness. The hUCMSC-EVs enriched with these two miRNAs significantly suppressed LFH in vivo and notably ameliorated the progression of transforming growth factor ß1(TGF-ß1)-induced fibrosis in vitro after delivering these two miRNAs to mouse LF cells. The results further demonstrated that miR-146a-5p and miR-221-3p directly bonded to the 3'-UTR regions of SMAD4 mRNA, thereby inhibiting the TGF-ß/SMAD4 signalling pathway. Therefore, this translational study determined the effectiveness of a hUCMSC-EVs-based approach for the treatment of LFH and revealed the critical target of miR-146a-5p and miR-221-3p. Our findings provide new insights into promising therapeutics using a hUCMSC-EVs-based delivery system for patients with lumbar spinal stenosis.

Endo-TLIF versus MIS-TLIF in 1-segment lumbar spondylolisthesis: A prospective randomized pilot study.

OBJECTIVE: To evaluate the curative efficacy by comparing perioperative characteristics and 1.5-year observational outcomes in 1-segment lumbar spondylolisthesis between traditional minimally invasive transforaminal lumbar interbody fusion (MIS-TLIF) and optimized Endoscopic TLIF techniques. METHODS: The study was a single-center, randomized controlled trial comparing two different treatment approaches for 1-segment lumbar spondylolisthesis. 102 patients treated by MIS-TLIF (48 cases) or Endo-TLIF (54 cases) were included from March 2018 to April 2019. Perioperative parameters and clinical outcomes were evaluated. Degree of slip were measured, and fusion rates were determined at 18 months after surgery. RESULTS: The Endo-TLIF group had similar return to work time and rate. Blood loss, left bed time, analgesic ratio were significantly less in Endo-TLIF group. The Endo-TLIF group had a significantly longer operative time. Significant postoperative reduction in %slip was showed in both groups. The VAS and ODI improved significantly in both groups after surgery. Significant decreases in low-back pain in Endo-TLIF group were found at postoperative day 1 and 3 months. The fusion rate in the two groups was similar. CONCLUSION: Endo-TLIF surgery with a C-shaped working tube and a visualization system may be regarded as an efficient alternative surgery for 1-segment lumbar spondylolisthesis. It is a safe and minimally invasive way to perform this surgery and has shown satisfactory clinical outcomes. TRIAL REGISTRATION: ChiCTR1800015197, 13 March 2018. TRIAL REGISTRY: Chinese Clinical Trial Registry. Registered 13 March 2018. http://www.chictr.org.cn/showproj.aspx?proj=25865.

Urolithin a alleviates oxidative stress-induced senescence in nucleus pulposus-derived mesenchymal stem cells through SIRT1/PGC-1α pathway.

BACKGROUND: In degenerative intervertebral disc (IVD), an unfavorable IVD environment leads to increased senescence of nucleus pulposus (NP)-derived mesenchymal stem cells (NPMSCs) and the inability to complete the differentiation from NPMSCs to NP cells, leading to further aggravation of IVD degeneration (IDD). Urolithin A (UA) has been proven to have obvious effects in delaying cell senescence and resisting oxidative stress. AIM: To explore whether UA can alleviate NPMSCs senescence and to elucidate the underlying mechanism. METHODS: In vitro, we harvested NPMSCs from rat tails, and divided NPMSCs into four groups: the control group, H2O2 group, H2O2 + UA group, and H2O2 + UA + SR-18292 group. Senescence-associated ß-Galactosidase (SA-ß-Gal) activity, cell cycle, cell proliferation ability, and the expression of senescence-related and silent information regulator of transcription 1/PPAR gamma coactivator-1α (SIRT1/ PGC-1α) pathway-related proteins and mRNA were used to evaluate the protective effects of UA. In vivo, an animal model of IDD was constructed, and X-rays, magnetic resonance imaging, and histological analysis were used to assess whether UA could alleviate IDD in vivo. RESULTS: We found that H2O2 can cause NPMSCs senescence changes, such as cell cycle arrest, reduced cell proliferation ability, increased SA-ß-Gal activity, and increased expression of senescence-related proteins and mRNA. After UA pretreatment, the abovementioned senescence indicators were significantly alleviated. To further demonstrate the mechanism of UA, we evaluated the mitochondrial membrane potential and the SIRT1/PGC-1α pathway that regulates mitochondrial function. UA protected mitochondrial function and delayed NPMSCs senescence by activating the SIRT1/PGC-1α pathway. In vivo, we found that UA treatment alleviated an animal model of IDD by assessing the disc height index, Pfirrmann grade and the histological score. CONCLUSION: In summary, UA could activate the SIRT1/PGC-1α signaling pathway to protect mitochondrial function and alleviate cell senescence and IDD in vivo and vitro.

Sulfatinib, a novel kinase inhibitor, in patients with advanced solid tumors: results from a phase I study.

Sulfatinib is a small molecule kinase inhibitor that targets tumor angiogenesis and immune modulation. This phase I study (NCT02133157) investigated the safety, pharmacokinetic characteristics, and preliminary anti-tumor activity of sulfatinib in patients with advanced solid tumors. The study included a dose-escalation phase (50-350 mg/day, 28-day cycle) with a Fibonacci (3+3) design, and a tumor-specific expansion phase investigating the tumor response to treatment. Two sulfatinib formulations were assessed: formulation 1 (5, 25, and 50 mg capsules) and formulation 2 (50 and 200 mg capsules). Seventy-seven Chinese patients received oral sulfatinib; the maximum tolerated dose was not reached. Dose-limiting toxicities included abnormal hepatic function and coagulation tests, and upper gastrointestinal hemorrhage. The most common treatment-related adverse events were proteinuria, hypertension and diarrhea. Among 34 patients receiving sulfatinib formulation 2, one patient with hepatocellular carcinoma and eight with neuroendocrine tumors exhibited a partial response; 15 had stable disease. The objective response rate was 26.5% (9/34) and the disease control rate was 70.6% (24/34). Pharmacokinetic, safety, and efficacy data supported continuous oral administration of sulfatinib at 300 mg as the recommended phase II dose. Sulfatinib exhibited an acceptable safety profile and encouraging antitumor activity in patients with advanced solid tumors, particularly neuroendocrine tumors.

MRI assessment of lumbar intervertebral disc degeneration with lumbar degenerative disease using the Pfirrmann grading systems.

BACKGROUND: To evaluate by MRI intervertebral disc degeneration in patients with lumbar degenerative disease using the Pfirrmann grading system and to determine whether Modic changes correlated with the Pfirrmann grades and modified Pfirrmann grades of disc degeneration. METHODS: The clinical data of 108 surgical patients with lumbar degenerative disease were reviewed and their preoperative MR images were analyzed. Disc degeneration was evaluated using the Pfirrmann grading system. Patients were followed up and low back pain was evaluated using the visual analog scale (VAS) and the effect of back pain on the daily quality of life was assessed using Oswestry disability index (ODI). RESULTS: Forty-four cases had normal anatomical appearance (Modic type 0) and their Pfirrmann grades were 3.77±0.480 and their modified Pfirrmann grades were of 5.81±1.006. Twenty-seven cases had Modic type I changes and their Pfirrmann grades were 4.79±0.557 and their modified Pfirrmann grades were 7.00±0.832. Thirty-six cases exhibited Modic type II changes and their Pfirrmann grades and modified Pfirrmann grades were 4.11±0.398 and 6.64±0.867, respectively. One case had Modic type III changes. Kruskal-Wallis test revealed significant difference in modified Pfirrmann grade among Modic type 0, I and II changes (P<0.01) but no significant difference between Modic type I and II changes (P>0.05). Binary regression analysis showed that Modic changes correlated most strongly with disc degeneration. Follow up studies indicated that the VAS and ODI scores were markedly improved postoperatively. However, no difference was noted in VAS and ODI scores among patients with different Modic types. CONCLUSION: Modic changes correlate with the Pfirrmann and modified Pfirrmann grades of disc degeneration in lumbar degenerative disease. There is no significant correlation between Modic types and surgical outcomes.

[Detecting the information of cucumber in greenhouse for picking based on NIR image].

For the cucumber harvesting robot, the identification of target information is one of important tasks in the automation of fruit-picking. In order to implement spatial fruit localization and quality discrimination in greenhouse, this paper presented a machine vision algorithm for the recognition and detection of cucumber fruits based on near-infrared spectral imaging. By comparing the spectral reflectance of cucumber plant (fruit, leaf and stem) from visible to infrared region (325-1 075 nm) measured by ASD FieldSpec Pro VNIR spectrometer, a monospectral near-infrared image at the 850 nm sensitive wavelength was captured to cope with the similar-color segmentation problem in complex environment. Then, a method of fruit extraction was developed on the basis of the following steps. Firstly, from the gray level histogram it was observed that the pixels of fruit distributed on the right are lesser than that of background, so "P parameter threshold method" was used to image segmentation. Subsequently, divided local image was partitioned into several sub-blocks by the application of adaptive template mining, which was feasible for processing the fruit with long-column feature. Finally, noises including parts of stem and leaf were eliminated using estimation condition of barycentre position and area size, proved by relative experiment In addition, the region for robotic grasping was established by gray variation between fruit-handle and fruit pedicel, as the quality feature was extracted with morphological characteristics of the centre-line length and the fruit flexure degree. A detecting experiment was carried out on 30 images with cucumber fruits and 10 images with no fruits, which were taken in a changing greenhouse environment. The results indicate that the accuracy rate of the recognition was 83.3% and 100%, while the success rate of effectively acquiring the grasping region was 83.3%, which can meet the demand of robotic fruit-harvesting.

Myeloid suppressor cell-associated immune dysfunction in CSA1M fibrosarcoma tumor-bearing mice.

CSA1M tumor-bearing mice exhibited a severe immune dysfunction but the underlying mechanism remained unclear. In this study, we demonstrated that the myeloid suppressor cell (Mac-1(+)Gr-1(+) cells)-(MSC) related T cell immunosuppression in this tumor-bearing model. In mice at the late stage of CSA1M tumor-bearing (Late TB [8-10 weeks after cell inoculation in male BALB/c mice]), the percentages for CD4(+) and CD8(+) T cells decreased but Mac-1(+) cells increased in spleens with severe splenomegaly. There was no deficit for concanavalin A-induced CD4(+) and CD8(+) T cell proliferation, interferon-gamma (IFN-gamma) and interleukin (IL)-4 production, but delayed-type hypersensitivity reaction were attenuated. Analysis of cytokine production in unfractionated spleen cells showed a significant reduction of IFN-gamma and a marked increase of IL-10 and IL-4. In Late-TB mice, splenic MSC number intensively accumulated; the mRNA expressions of the signal transducer and activator of transcription 1, interferon regulatory factor 1 (IRF-1), and inducible nitric-oxide synthase (iNOS) were enhanced in MSC; the nitric oxide production and arginase enzyme activity increased in MSC as well. Furthermore, the concanavalin A-induced T cell proliferation was inhibited in the presence of lipopolysaccharide- or IFN-gamma-activated MSC from Late-TB mice, which could be reversed by the iNOS specific inhibitor L-NMMA. iNOS seemed to be required more than arginase for the suppressive activity of MSC. Taken together, our results suggest that the immune dysfunction in tumor-bearing mice might be causally associated with the accumulation of MSC and its tumor-favoring property.

(5R)-5-hydroxytriptolide (LLDT-8) protects against bleomycin-induced lung fibrosis in mice.

AIM: To study the protective effects of a triptolide-derived, novel compound, (5R)-5-hydroxytriptolide (LLDT-8), on bleomycin-induced lung fibrosis. METHODS: C57BL/6 mice received an intratracheal injection of bleomycin and were then treated with LLDT-8 (0.5, 1, 2 mg/kg, ip) once daily for 7 or 14 consecutive days. The body weight loss and lung index augmentation was observed; the inflammatory response including differential cells counts of neutrophils, macrophages, and lymphocytes in the bronchoalveolar lavage fluid (BALF), superoxide dismutase (SOD), and malondialdehyde (MDA) level in the lung homogenates was detected, and the fibrosis extent was evaluated by hydroxyproline content and histopathological changes in the lungs. In addition, the pro-inflammatory and pro-fibrotic cytokines, tumor necrosis factor-alpha (TNF-alpha), interleukin-4 (IL-4), and transforming growth factor-alpha (TGF-alpha) production in the lungs were measured. RESULTS: LLDT-8 alleviated the body weight loss and lung index increase caused by bleomycin, reduced neutrophils and lymphocytes in the BALF, promoted SOD activity, decreased MDA production, and inhibited the hydroxyproline level and the amelioration of lung tissue histological damage. Moreover, LLDT-8 suppressed TNF-alpha, IL-4, and TGF-beta production in the lung homogenates. CONCLUSION: LLDT-8 showed protective effects against bleomycin-induced lung fibrosis, and the results suggested the potential role of LLDT-8 in the treatment of this disease.

Preventive effects of (5R)-5-hydroxytriptolide on concanavalin A-induced hepatitis.

(5R)-5-hydroxytriptolide (LLDT-8) exhibits strong immunosuppressive activities in vitro and in vivo. Here, we investigated the effects of LLDT-8 on concanavalin A-induced hepatitis. Liver damage was evaluated by serum alanine transaminase (ALT) level and liver histology. The effects of LLDT-8 were determined by measurement of serum cytokines, lymphocyte proliferation assay, flow cytometry analysis of splenic T cell percentage and apoptosis, reverse-transcription polymerase chain reaction (RT-PCR) analysis for gene transcriptions. In LLDT-8-treated mice, serum ALT level and histological damage were markedly attenuated. The beneficial effect of LLDT-8 was closely associated with (i) reduction of serum tumor necrosis factor-alpha, interferon-gamma (IFN-gamma), interleukin-2, interleukin-12, and interleukin-6 levels; (ii) elimination of activated T cells by increasing proapoptotic genes signal transducer and activator of transcription 1 (STAT1) and interferon regulatory factor-1 (IRF-1) expression in spleens; (iii) blockade of mRNA expressions for chemokines (monokine induced by IFN-gamma, Mig; IFN-gamma-inducible protein-10, IP-10; IFN-inducible T cell-alpha chemoattractant, I-TAC), vascular adhesion molecule-1 (VCAM-1), and chemokine receptors (C-C chemokine receptor 1, CCR1; C-C chemokine receptor 5, CCR5; C-X-C chemokine receptor 3, CXCR3) in livers. These results suggested the therapeutic potential of LLDT-8 in IFN-gamma/STAT1/IRF-1 signaling- and inflammatory cytokines-mediated immune disorders.

Differential expression of inducible nitric oxide synthase and IL-12 between peritoneal and splenic macrophages stimulated with LPS plus IFN-gamma is associated with the activation of extracellular signal-related kinase.

Resident peritoneal macrophages (pMphi) are found deficient in T cell-stimulating capacity compared with the competent splenic macrophages (sMphi). Macrophages (Mphi)-derived nitric oxide (NO) and IL-12 have been shown to play crucial roles in the interaction between Mphi and T cells. To further understand differential functions between pMphi and sMphi, we focused on the production of NO and IL-12 from LPS plus IFN-gamma-activated Mphi. We demonstrated the differential expression of inducible nitric oxide synthase (iNOS) and IL-12 in pMphi and sMphi with LPS plus IFN-gamma stimulation. pMphi produced high level of NO but low level of IL-12, whereas sMphi produced high level of IL-12 but no NO. Furthermore, we demonstrated that there were no differences in IFN-gamma-induced signal transducer and activator of transcription-1 activation and consequent interferon regulatory factor-1 and interferon consensus sequence-binding protein up-regulation between pMphi and sMphi. Likewise, p38 mitogen-activated protein kinase was activated by LPS with identical kinetics in both pMphi and sMphi. However, LPS-induced extracellular signal-regulated kinase (ERK) activation was prolonged in pMphi comparing with sMphi. Moreover, we demonstrated, using inhibitor selective for ERK cascade (PD98059), that the prolonged ERK activation contributed a positive signal for iNOS expression and a negative signal for IL-12p40 expression in resident pMphi. In addition, anti-IL-10-neutralizing antibody plus indomethacin could abrogate the inhibitory effects of endogenous IL-10 and prostaglandin E2 on the production of IL-12 by resident pMphi possibly through suppressing ERK activation. Taken together, profound difference in ERK activation may account for differential LPS plus IFN-gamma responsiveness between pMphi and sMphi. High production of NO and low production of IL-12 by pMphi may contribute to its deficiency in T cell-stimulating capacity.

(5R)-5-hydroxytriptolide attenuated collagen-induced arthritis in DBA/1 mice via suppressing interferon-gamma production and its related signaling.

(5R)-5-Hydroxytriptolide (LLDT-8) displays strong immunosuppressive activities both in vitro and in vivo in our previous studies. This study aims to investigate whether LLDT-8 has antiarthritic potential in a murine model of type II bovine collagen (CII)-induced arthritis (CIA) and to show the mechanism(s) of LLDT-8 action. DBA/1 mice were immunized with CII to induce arthritis and administered with LLDT-8. The severity of arthritis was evaluated according to the clinical score and joint damage. The effects of LLDT-8 on immune responses were determined by measurement of serum antibody levels, lymphocyte proliferation assay, cytokine assay, nitric oxide (NO) production, arginase activity assays, fluorescence-activated cell sorting analysis of splenic Mac-1+ cells, as well as polymerase chain reaction analysis for interferon-gamma (IFN-gamma)-related gene expression. We showed that LLDT-8 treatment significantly reduced the incidence and severity of CIA. The preventive and therapeutic effects of LLDT-8 are associated with 1) reduction of serum anti-CII immunoglobulin (Ig) G, IgG2a, and IgG1 levels; 2) inhibition of CII-specific lymphocyte proliferation, IFN-gamma and interleukin-2 production; 3) blockade of gene expressions in IFN-gamma signaling, including IFN-gamma production pathways [signal transducer and activator of transcription (STAT) 1, T-box transcription factor, interleukin 12Rbeta2, and STAT4] and IFN-gamma-induced chemokine transcription [macrophage inflammatory protein (Mip)-1alpha, Mip-1beta, regulated on activation normally T cell expressed and secreted, and inducible protein 10]; and 4) retardation of the abnormal increase of NO via IFN-gamma/STAT1/interferon regulatory factor 1/inducible nitric-oxide synthase pathway and arginase activity. Moreover, the mRNA transcription of chemokine receptors was also suppressed [including C-C chemokine receptor (CCR) 1, CCR5, and C-X-C chemokine receptor 3]. In conclusion, our data suggest that the antiarthritic effect of LLDT-8 is closely related to the blockade of IFN-gamma signaling. LLDT-8 may have a therapeutic value in the treatment of rheumatoid arthritis.

[Experimental study of modified bio-derived material by pluronic F-127 in vitro].

OBJECTIVE: To investigate the effects of three-dimentional culture in bioderived material modified by Pluronic F-127 on the growth and function of rabbit periosteal osteoblast in vitro. METHODS: Bio-derived materials were from fresh pig ribs and were modified by Pluronic F-127. Then rabbit periosteal osteoblasts were cultured in bio-derived materials(group A), in the modified bio-derived materials(group B) and on the plastic surfaces as a control (group C), respectively. During a 7-day period, the status of growth, cell viability and alkaline phosphatase(ALP) activity were measured. RESULTS: Osteoblasts attached, elongated and grew well on the modified bio-derived materials. There were no significant difference in osteogenesis and ALP activity between group A and group B(P > 0.05). The osteogenesis and ALP activity in groups A, B were less than those in group C (P < 0.01). CONCLUSION: Pluronic F-127 can be used for a carrier for bioactive factors to modify bio-derived material.

[An experimental study on effect of estrogen on osteoarthritis in female rats].

OBJECTIVE: To investigate the effect of estrogen on osteoarthritis in female rats. METHODS: Forty female rats were divided into four groups. In group I, the rats were not given any treatment as a control. In groups II, III and IV, the rats received fixing left knee joint on extension position. Meanwhile, the rats received ovariectomy in group III; ovariectomy and diethylstilbestrol treatment in group IV, respectively. After 4 weeks, histological observation and serum BGP examination were done. RESULTS: In groups II, III and IV, the levels of serum BGP were 3.50 +/- 0.39, 5.72 +/- 0.64 and 3.95 +/- 0.44, respectively. The pathologic grades of cartilage and synovium were 10.83 +/- 4.35 and 4.21 +/- 2.03; 15.32 +/- 3.42 and 7.62 +/- 3.42; and 12.65 +/- 2.73 and 5.46 +/- 1.23, respectively. CONCLUSION: Estrogen may play an important role in delaying the development of osteoarthritis.