Tibial Cortex Transverse Transport Facilitates Severe Diabetic Foot Wound Healing via HIF-1α-Induced Angiogenesis.

Purpose: Management of severe diabetic foot ulcers (DFUs) remains challenging. Tibial cortex transverse transport (TTT) facilitates healing and limb salvage in patients with recalcitrant DFUs. However, the underlying mechanism is largely unknown, necessitating the establishment of an animal model and mechanism exploration. Methods: Severe DFUs were induced in rats, then assigned to TTT, sham, or control groups (n=16/group). The TTT group underwent a tibial corticotomy, with 6 days each of medial and lateral transport; the sham group had a corticotomy without transport. Ulcer healing was assessed through Laser Doppler, CT angiography, histology, and immunohistochemistry. Serum HIF-1α, PDGF-BB, SDF-1, and VEGF levels were measured by ELISA. Results: The TTT group showed lower percentages of wound area, higher dermis thickness (all p < 0.001 expect for p = 0.001 for TTT vs Sham at day 6) and percentage of collagen content (all p < 0.001) than the other two groups. The TTT group had higher perfusion and vessel volume in the hindlimb (all p < 0.001). The number of CD31+ cells (all p < 0.001) and VEGFR2+ cells (at day 6, TTT vs Control, p = 0.001, TTT vs Sham, p = 0.006; at day 12, TTT vs Control, p = 0.003, TTT vs Sham, p = 0.01) were higher in the TTT group. The activity of HIF-1α, PDGF-BB, and SDF-1 was increased in the TTT group (all p < 0.001 except for SDF-1 at day 12, TTT vs Sham, p = 0.005). The TTT group had higher levels of HIF-1α, PDGF-BB, SDF-1, and VEGF in serum than the other groups (all p < 0.001). Conclusion: TTT enhanced neovascularization and perfusion at the hindlimb and accelerated healing of the severe DFUs. The underlying mechanism is related to HIF-1α-induced angiogenesis.

Tibial cortex transverse transport promotes ischemic diabetic foot ulcer healing via enhanced angiogenesis and inflammation modulation in a novel rat model.

BACKGROUND: Tibial Cortex Transverse Transport (TTT) represents an innovative surgical method for treating lower extremity diabetic foot ulcers (DFUs), yet its underlying mechanisms remain elusive. Establishing an animal model that closely mirrors clinical scenarios is both critical and novel for elucidating the mechanisms of TTT. METHODS: We established a diabetic rat model with induced hindlimb ischemia to mimic the clinical manifestation of DFUs. TTT was applied using an external fixator for regulated bone movement. Treatment efficacy was evaluated through wound healing assessments, histological analyses, and immunohistochemical techniques to elucidate biological processes. RESULTS: The TTT group demonstrated expedited wound healing, improved skin tissue regeneration, and diminished inflammation relative to controls. Marked neovascularization and upregulation of angiogenic factors were observed, with the HIF-1α/SDF-1/CXCR4 pathway and an increase in EPCs being pivotal in these processes. A transition toward anti-inflammatory M2 macrophages indicated TTT's immunomodulatory capacity. CONCLUSION: Our innovative rat model effectively demonstrates the therapeutic potential of TTT in treating DFUs. We identified TTT's roles in promoting angiogenesis and modulating the immune system. This paves the way for further in-depth research and potential clinical applications to improve DFU management strategies.

Efficacy and safety of unilateral tibial cortex transverse transport on bilateral diabetic foot ulcers: A propensity score matching study.

Background: Tibial Cortex Transverse Transport (TTT) has been demonstrated to be an effective treatment for unilateral diabetic foot ulcers (UDFUs). However, this retrospective study was designed to compare the efficacy and safety of unilateral TTT on bilateral diabetic foot ulcers (BDFUs). Methods: This retrospective study included a review of patients with TTT treated from January 2017 to August 2019, Propensity Score Matching (PSM) was performed to compare patients with BDFUs to those with UDFUs. Ulcer healing, recurrence, and major amputation rates were evaluated at 1-year follow-up. Changes in foot vessels were assessed in the BDFUs group using computed tomography angiography (CTA). Results: A total of 140 patients with DFUs (106 UDFUs and 34 BDFUs) were included in the study. UDFUs and BDFUs were matched in a 1:1 ratio (34 in each group) using PSM. No significant difference was observed at 1-year-follow-up [91.2% (31/34) vs. 76.5% (26/34), OR 0.315 (95% CI 0.08 to 1.31), P â€‹= â€‹0.10] and 6-month-follow-up [70.6% (24/34) vs. 50.0% (17/34), OR 0.85 (95% CI 0.15 to 1.13), P â€‹= â€‹0.08] in two groups. Significant differences in rates of major amputation and recurrence between the groups (P â€‹> â€‹0.05) were not observed. The BDFUs group appeared more angiogenesis of the foot by CTA after 8 weeks of operation. Conclusion: Results of this study suggest that severe BDFUs can be effectively treated by unilateral TTT. TTT is easy to operate and effective, which may be a good alternative for treating severe BDFUs. The translational potential of this article: In previous retrospective clinical studies, TTT has demonstrated promising clinical outcomes in the management of diabetic foot ulcers. In this current study, we aim to investigate the potential use of TTT in treating distant tissue defects by evaluating the limited availability and safety of TTT for the management of bilateral diabetic foot. While additional basic and clinical research is necessary to fully elucidate the underlying mechanisms, our study offers insight into the potential therapeutic use of TTT for this condition.

Effect of tibial cortex transverse transport in patients with recalcitrant diabetic foot ulcers: A prospective multicenter cohort study.

Background: Management of recalcitrant diabetic foot ulcer (DFU) remains difficult. Distraction osteogenesis mediates new bone formation and angiogenesis in the bone itself and the surrounding tissues. Recently it was reported that tibial cortex transverse transport (TTT) was associated with neovascularization and increased perfusion at the foot in patients with recalcitrant DFUs and facilitated healing and limb salvage. However, the findings were from several single-center studies with relatively small populations, which need to be confirmed in multicenter cohort studies with relatively large populations. Furthermore, the effect of this technique on patient's health-related quality of life is still unclear. Methods: We treated patients with recalcitrant (University of Texas wound grading system 2-C to 3-D and not responding to prior routine conservative and surgical treatments for at least 8 weeks) DFUs from seven centers using TTT (a 5 â€‹cm â€‹× â€‹1.5 â€‹cm corticotomy followed by 4 weeks of medial and lateral distraction) between July 2016 and June 2019. We analyzed ulcer healing, major amputation, recurrence, health-related quality of life (physical and mental component summary scores), and complications in the 2-year follow-up. Foot arterial and perfusion changes were evaluated using computed tomography angiography and perfusion imaging 12 weeks postoperatively. Results: A total of 1175 patients were enrolled. Patients who died (85, 7.2%) or lost to follow-up (18, 1.7%) were excluded, leaving 1072 patients for evaluation. Most of the patients were male (752, 70.1%) and with a mean age of 60.4 â€‹± â€‹9.1 years. The mean ulcer size was 41.0 â€‹± â€‹8.5 â€‹cm2 and 187 (16.6%) ulcers extended above the ankle. During the follow-up, 1019 (94.9%) patients healed in a mean time of 12.4 â€‹± â€‹5.6 weeks, 53 (4.9%) had major amputations, and 33 (3.1%) experienced recurrences. Compared to preoperatively, the patients had higher physical (26.2 â€‹± â€‹8.3 versus 41.3 â€‹± â€‹10.6, p â€‹= â€‹0.008) and mental (33.6 â€‹± â€‹10.7 versus 45.4 â€‹± â€‹11.3, p â€‹= â€‹0.031) component summary scores at the 2-year follow-up. Closed tibial fracture at the corticotomy site was found in 8 (0.7%) patients and was treated using external fixation and healed uneventfully. There were 23 (2.1%) patients who had pin site infections and were treated successfully with dressing changes. Compared to preoperatively, the patients had more small arteries and higher foot blood flow (8.1 â€‹± â€‹2.2 versus 28.3 â€‹± â€‹3.9 ml/100 â€‹g/min, p â€‹= â€‹0.003) and volume (1.5 â€‹± â€‹0.3 versus 2.7 â€‹± â€‹0.4 ml/100 â€‹g, p â€‹= â€‹0.037) 12 weeks postoperatively. Conclusion: TTT promotes healing, limb salvage, and health-related quality of life in patients with recalcitrant DFUs as demonstrated in this multicenter cohort study. The surgical procedure was simple and straightforward and the complications were few and minor. The effect of this technique was associated with neovascularization and improved perfusion at the foot mediated by the cortex distraction. The findings are required to confirm in randomized controlled trials.The Translational Potential of this Article: TTT can be used as an effective treatment in patients with recalcitrant DFUs. The mechanism is associated with neovascularization and consequently increased perfusion in the foot after operation.

Knowledge Domains and Emerging Trends of Osteoblasts-Osteoclasts in Bone Disease From 2002 to 2021: A Bibliometrics Analysis and Visualization Study.

Background: Osteoblasts-Osteoclasts has been a major area in bone disease research for a long time. However, there are few systematic studies in this field using bibliometric analysis. We aimed to perform a bibliometric analysis and visualization study to determine hotspots and trends of osteoblasts-osteoclasts in bone diseases, identify collaboration and influence among authors, countries, institutions, and journals, and assess the knowledge base to develop basic and clinical research in the future. Methods: We collected articles and reviews for osteoblasts-osteoclasts in bone diseases from the Web of Science Core Collection. In addition, we utilized scientometrics software (CiteSpace5.8 and VOSviewer1.6.18) for visual analysis of countries/regions, institutions, authors, references, and keywords in the field. Results: In total, 16,832 authors from 579 institutions in 73 countries/regions have published 3,490 papers in 928 academic journals. The literature in this field is rapidly increasing, with Bone publishing the most articles, whereas Journal of Bone and Mineral Research had the most co-cited journals. These two journals mainly focused on molecular biology and the clinical medicine domain. The countries with the highest number of publications were the US and China, and the University of Arkansas for Medical Sciences was the most active institution. Regarding authors, Stavros C. Manolagas published the most articles, and Hiroshi Takayanagi had the most co-cited papers. Research in this field mainly includes molecular expression and regulatory mechanisms, differentiation, osteoprotection, inflammation, and tumors. The latest research hotspots are oxidative stress, mutation, osteocyte formation and absorption, bone metabolism, tumor therapy, and in-depth mechanisms. Conclusion: We identified the research hotspots and development process of osteoblasts-osteoclasts in bone disease using bibliometric and visual methods. Osteoblasts-osteoclasts have attracted increasing attention in bone disease. This study will provide a valuable reference for researchers concerned with osteoblasts-osteoclasts in bone diseases.

[Treatment of severe diabetic foot ulcer using tibia transverse transport combined with nose ring drain].

OBJECTIVE: To investigate the effectiveness of tibial transverse transport (TTT) combined with nose ring drain (NRD) in the treatment of severe diabetic foot ulcer. METHODS: The clinical data of 60 patients with severe diabetic foot (Wagner grade 3 or 4) ulcer who were admitted between April 2017 and August 2020 and met the selection criteria were retrospectively analyzed. Among them, 30 cases were treated with TTT combined with NRD (TTT+NRD group), and 30 cases were treated with TTT (TTT group). There was no significant difference in gender, age, diabetes duration, preoperative glycated hemoglobin, comorbidities, wound area, and duration, side, and grade of diabetic foot ( P>0.05). The wound healing time, wound healing rate, amputation rate, recurrence rate, duration of antibiotic therapy, hospital stay, number of hospitalizations, and number of operations were recoreded and compared between the two groups. RESULTS: No obvious surgical complications occurred in the two groups. Patients in both groups were followed up 3-13 months, with an average of 5.7 months. The duration of antibiotic therapy and hospital stay in the TTT+NRD group were significantly shorter than those in the TTT group ( P<0.05). There was no significant difference in wound healing time, wound healing rate, number of hospitalizations, and number of operations between the two groups ( P>0.05). During follow-up, there was no recurrence of ulcer in the TTT+NRD group while 2 recurrent cases (6.7%) in the TTT group. The difference in recurrence rate was not significant ( P=0.492). One case (3.3%) in the TTT+NRD group underwent amputation due to acute lower extremity vascular embolism, and 1 case (3.3%) in the TTT group underwent amputation due to secondary necrosis. The difference in amputation rate was not significant between the two groups ( P=1.000). CONCLUSION: TTT combined with NRD is an effective method for the treatment of severe diabetic foot ulcers with deep infections or relatively closed cavities or sinuses. It can shorten the time of antibiotic use and the length of hospitalization; and the NRD has a good drainage effect without obvious comorbidities, procedure and the postoperative care are simple and easy to obtain materials.

Tibial cortex transverse transport facilitating healing in patients with recalcitrant non-diabetic leg ulcers.

OBJECTIVE: The treatment of recalcitrant not-diabetic leg ulcers remains challenging. Distraction osteogenesis is accompanying by angiogenesis and neovascularization in the surrounding tissues. We previously applied tibial cortex transverse transport (TTT) to patients with recalcitrant diabetic foot ulcers and found neovascularization and increased perfusion in the foot and consequently enhanced healing and limb salvage and reduced recurrence. However, the effects of TTT on recalcitrant non-diabetic leg ulcer remains largely unknown. METHODS: Consecutive patients (n â€‹= â€‹85) with recalcitrant non-diabetic leg ulcers (University of Texas Grade 2-B to 3-D, ie, wound penetrating to the tendon, capsule, bone, or joint with infection and/or ischemia) were recruited and divided into TTT (n â€‹= â€‹42) and control (n â€‹= â€‹43) groups based on the treatment they received. There were 36 (85.7%) arterial ulcers, 4 (9.5%) venous ulcers and 2 (4.8%) mixed ulcers in the TTT group and 32 (74.4%) arterial ulcers, 7 (16.7%) venous ulcers and 4 (9.3%) mixed ulcers in the control group (p â€‹> â€‹0.05). The two groups were matched on demographic and clinical characteristics. Patients in the TTT group underwent tibial corticotomy followed by 4 weeks of distraction medially then laterally, while those in the control group received conventional surgeries (debridements, revascularization, reconstruction with flaps, or skin grafts or equivalents). Ulcer healing and healing time, limb salvage, recurrence, and patient death were evaluated at a 1-year follow-up. Changes in leg small vessels were assessed in the TTT group using computed tomography angiography (CTA). RESULTS: TTT group had higher healing rates at 1-year follow-up than the control group (78.6% [33/42] vs. 58.1% [25/43], OR 2.64 [95% CI 1.10 to 6.85], p â€‹= â€‹0.04). The healing time of the TTT group was shorter than the control group (4.5 vs. 6.1 months, mean difference -1.60 [95% CI -2.93 to -0.26], p â€‹= â€‹0.02). There were no significant differences in rates of major amputation, reulceration, or mortality between the groups (p â€‹> â€‹0.05). TTT group displayed more small vessels 4 weeks postoperatively at the wound area, the foot, and the calf of the ipsilateral side in CTA. All patients in the TTT group achieved good union at the osteotomy site and had no skin or soft tissue necrosis or infection around the incision area. CONCLUSION: The findings showed that TTT facilitated the healing of recalcitrant non-diabetic leg ulcers and reduced the healing time compared with conventional surgeries. They suggest that TTT is an effective procedure to treat recalcitrant non-diabetic foot ulcers compared with standard surgical therapy. The procedure of TTT is relatively simple. Randomized controlled trials are required to confirm these findings. THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE: TTT can be used as an effective treatment for recalcitrant non-diabetic leg ulcers in patients. The mechanism may be associated with the neovascularization in the ulcerated foot induced by TTT and consequently increased perfusion. Together with previous findings from recalcitrant diabetic leg ulcers, the findings suggest TTT as an effective procedure to treat recalcitrant chronic leg ulcers.

[Treatment experiences of 516 cases of diabetic foot treated with tibial transverse transport].

Tibial transverse transport (TTT) was firstly applied to treat thromboangiitis obliterans successfully by Professor QU Long in China in 2000. Based on this, the team of Professor HUA Qikai in the First Affiliated Hospital of Guangxi Medical University applied this technique to treat diabetic foot since 2013, and until now, more than 500 patients underwent this treatment with excellent effectiveness including a salvage rate as high as 96.1%. Our team also improved this technique in many aspects, and developed a TTT-based classification system and treatment for diabetic foot. We also explored the underlying mechanism of TTT treatment using imaging, histology, and other basic research methods. To further promote the application of this technique in clinic, we reported the findings from our cases and reviewed our previous findings in this study.

Proximal Tibial Cortex Transverse Distraction Facilitating Healing and Limb Salvage in Severe and Recalcitrant Diabetic Foot Ulcers.

BACKGROUND: The management of severe and recalcitrant diabetic foot ulcers is challenging. Distraction osteogenesis is accompanied by vascularization and regeneration of the surrounding tissues. Longitudinal distraction of the proximal tibia stimulates increased and prolonged blood flow to the distal tibia. However, the effects of transverse distraction of the proximal tibia cortex on severe and recalcitrant diabetic foot ulcers are largely unknown. QUESTIONS/PURPOSES: (1) Does tibial cortex transverse distraction increase healing and decrease major amputation and recurrence of severe and recalcitrant diabetic foot ulcers compared with routine management (which generally included débridement, revascularization, negative pressure wound therapy, local or free flaps, or skin grafts as indicated)? (2) Does neovascularization and perfusion increase at the foot after the procedure? (3) What are the complications of tibial cortex transverse distraction in patients with severe and recalcitrant diabetic foot ulcers? METHODS: Between July 2014 and March 2017, we treated 136 patients with diabetes mellitus and University of Texas Grade 2B to 3D ulcers (wound penetrating to the tendon, capsule, bone, or joint with infection and/or ischemia). The patients had failed to respond to treatment for at least 6 months, and their ulcers had a mean ± SD area of 44 cm ± 10 cm. All 136 patients underwent tibial cortex transverse distraction (partial corticotomy of the upper tibia, which was in normal condition, followed by 4 weeks of transverse distraction medially then laterally). We compared these patients with the last 137 consecutive patients we treated with standard surgical treatment, consisting of débridement, revascularization, local or free flap or skin equivalent, or graft reconstruction along with negative-pressure wound therapy between May 2011 and June 2013; there was a 1-year period during which both treatments were in use, and we did not include patients whose procedures were performed during this time in either group. Patients in both groups received standard off-loading and wound care. The patients lost to follow-up by 2 years (0.7% of the treatment group [one of 137] and 1.4% of the control group [two of 139]; p = 0.57) were excluded. The patients in the treatment and control groups had a mean age of 61 years and 60 years, respectively, and they were predominantly men in both groups (70% [95 of 136] versus 64% [88 of 137]; p = 0.32). There were no differences with respect to parameters associated with the likelihood of ulcer healing, such as diabetes and ulcer duration, ulcer grades and area, smoking, and arterial status. We compared the groups with respect to ulcer healing (complete epithelialization without discharge, maintained for at least 2 weeks, which was determined by an assessor not involved with clinical care) in a 2-year follow-up, the proportion of ulcers that healed by 6 months, major amputation, recurrence, and complications in the 2-year follow-up. Foot arterial status and perfusion were assessed in the tibial cortex transverse distraction group using CT angiography and perfusion imaging. RESULTS: The tibial cortex transverse distraction group had a higher proportion of ulcers that healed in the 2-year follow-up than the control group (96% [131 of 136] versus 68% [98 of 137]; odds ratio 10.40 [95% confidence interval 3.96 to 27.43]; p < 0.001). By 6 months, a higher proportion of ulcers healed in the tibia cortex transverse distraction group than the control group (93% [126 of 136] versus 41% [56 of 137]; OR 18.2 [95% CI 8.80 to 37.76]; p < 0.001). Lower proportions of patients in the tibia cortex transverse distraction group underwent major amputation (2.9% [four of 136] versus 23% [31 of 137], OR 0.10 [95% CI 0.04 to 0.30]; p < 0.001) or had recurrences 2.9% (4 of 136) versus 17% (23 of 137), OR 0.20 [95% CI 0.05 to 0.45]; p < 0.001) than the control group in 2-year follow-up. In the feet of the patients in the tibial cortex transverse distraction group, there was a higher density of small vessels (19 ± 2.1/mm versus 9 ± 1.9/mm; mean difference 10/mm; p = 0.010), higher blood flow (24 ± 5 mL/100 g/min versus 8 ± 2.4 mL/100 g/min, mean difference 16 mL/100 g/min; p = 0.004) and blood volume (2.5 ± 0.29 mL/100 g versus 1.3 ± 0.33 mL/100 g, mean difference 1.2 mL/100 g; p = 0.03) 12 weeks postoperatively than preoperatively. Complications included closed fractures at the corticotomy site (in 1.5% of patients; two of 136), which were treated with closed reduction and healed, as well as pin-site infections (in 2.2% of patients; three of 136), which were treated with dressing changes and they resolved without osteomyelitis. CONCLUSIONS: Proximal tibial cortex transverse distraction substantially facilitated healing and limb salvage and decreased the recurrence of severe and recalcitrant diabetic foot ulcers. The surgical techniques were relatively straightforward although the treatment was unorthodox, and the complications were few and minor. These findings suggest that tibial cortex transverse distraction is an effective procedure to treat severe and recalcitrant diabetic foot ulcers compared with standard surgical therapy. Randomized controlled trials are required to confirm these findings. LEVEL OF EVIDENCE: Level II, therapeutic study.

Computed tomography perfusion study evaluating the curative effect of tibial transverse transport in patients with severe diabetic foot.

BACKGROUND: The clinical treatment of patients with severe diabetic foot (DF) is difficult. Recently, the First Affiliated Hospital of Guangxi Medical University began to apply tibial transverse transport (TTT) in patients with DF. This treatment has achieved significant effects, but its mechanism of action is unclear. Recently, microcirculation and the pathogenesis of diabetes have become the foci of research in this field. The evaluation of the possible mechanism of microcirculation reconstruction requires relevant indicators. The aim of this study was to investigate the value of computed tomography perfusion in evaluations of the curative effects of TTT and establish corresponding quantitative evaluation indicators. METHODS: Twelve patients with DF treated with TTT were recruited as the research participants. All diabetic feet were divided into the transport foot (TF) group and nontransport foot (NTF) group according to whether the patients underwent TTT. All patients underwent CT shuttle scanning preoperatively and 8 weeks after surgery. The shuttle scan data were transferred to Siemens VPCT body software and postprocessed with Customized Tumor2. We chose the TF posterior tibial artery from a distance of approximately 15 cm to the bifurcation of the plantar medial artery and the lateral plantar artery as the input artery. We selected the centre of the bilateral medial plantar muscle group on the coronal and axial regions of interest. We applied a deconvolution approach to obtain data from both sides of the plantar tissue perfusion. Skin temperature (ST) detection was performed with an ST gun to measure the average ST values in the dorsal and plantar areas, the first and fifth heads of the phalanges, and the medial and lateral malleolus points of both feet of patients with DF preoperatively and 8 weeks after surgery. RESULTS: The preoperative and postoperative ST values of the patients in the TF group were 30.73 ± 1.86 °C and 32.22 ± 1.51 °C, respectively. The preoperative and postoperative ST values for the patients in the NTF group were 30.93 ± 2.65 °C and 32.07 ± 2.09 °C, respectively. There were significant differences in the preoperative and postoperative data between the TF (P = 0.001) and NTF (P = 0.013) groups. In the patients with DF who underwent TTT, there were both preoperative and postoperative differences inside the medial plantar muscle group in the relative blood volume (rBV) value and relative mean transit time (rMTT) (P = 0.027, P = 0.026, respectively). The postoperative BV in the NTF group was increased compared with the preoperative BV (P = 0.006). CONCLUSION: There were significant differences in relative BV, relative mean transit time, and ST between the two groups before and after surgery, and the postoperative BV in the NTF group and ST values in the two groups were increased compared with the preoperative values. The BV in the NTF group and the ST values in the two groups were effective indicators in evaluating the changes between preoperative and postoperative perfusion. These results indicate that TTT could increase plantar tissue perfusion as assessed by BV and ST; this increase was among the reasons for the surface healing of severe DF ulcers. TRANSLATIONAL POTENTIAL OF THIS ARTICLE: Preoperative CT perfusion evaluation can provide relevant information of blood vessels and microcirculation for clinical operation, and postoperative CT perfusion evaluation can provide postoperative efficacy evaluation for clinical operation. All patients received information about the study and signed a specific informed consent. Approval for this study was granted by the regional ethics committee [Regional Ethics Committee of the First Affiliated Hospital of Guangxi Medical University, China [(2018-(KY-E-069].

[Apoptosis of hepatocellular induced by active ingredients of Zanthoxyli Radix by Raman spectroscopy].

The apoptosis of mono-hepatocellular induced by the active ingredients of the Zanthoxyli Radix was investigated using laser Raman spectroscopy. Hepatoma cells (BEL-7404) were treated with 10 mg•L⁻¹ nitidine chloride and 3 g•L⁻¹ the extracts of Zanthoxyli Radix, respectively, then were divided into two parts, one for fluorescence staining, the other for determination of Raman spectroscopy. The acquired spectra were then processed by background elimination, smoothing, and normalization. Fluorescence staining results showed that the nucleuses from untreated group were uniformly stained, while those from the group treated for 48 hours were densely stained and broken. The spectra results revealed that the intensity of peaks associated with nucleic acid and protein decreased after the cells were incubated with the extracts of Zanthoxyli Radix for 12, 24, 36 and 48 hours. The intensity of peaks at 785,1 002,1 175,1 660 cm⁻¹ was decreased with the time of the cells were incubated by the extracts of Zanthoxyli Radix. The results indicated that the extracts of Zanthoxyli Radix could induce the apoptosis of hepatoma cells and reduce the amount of nucleic acid and protein in the cells. There is a certain relevance between the drug treatment time and the efficacy. The above results suggest that Raman spectra can provide abundant information about the changes in biological macromolecules within the cells after incubated by the extracts of Zanthoxyli Radix and serve as an effective method for the real time measurement of apoptosis.

Mesenchymal stem cells suppress CD8+ T cell-mediated activation by suppressing natural killer group 2, member D protein receptor expression and secretion of prostaglandin E2, indoleamine 2, 3-dioxygenase and transforming growth factor-ß.

Bone marrow mesenchymal stem cells (BMSCs) inhibit immune cell responsiveness, and especially of T lymphocytes. We showed that BMSCs markedly inhibited the proliferation and cytokine production by CD8(+) T cells by a cell-to-cell contact phenomenon and secretion of soluble factors. BMSCs down-regulate the expression of natural killer group 2, member D protein (NKG2D) receptors on CD8(+) T cells when co-cultured with them. Moreover, CD8(+) T cells that express low levels of NKG2D had impaired proliferation after triggering by a mitogen. The major histocompatibility complex (MHC) class I chain-related (MIC) A/B molecule, which is a typical ligand for NKG2D, was expressed on BMSCs, and caused dampening of cell proliferation. Monoclonal antibody blocking experiments targeted to MIC A/B impaired CD8(+) T cell function, as evaluated by proliferation and cytokine production. In addition, the production of prostaglandin E2 (PGE2 ), indoleamine 2, 3-dioxygenase (IDO) and transforming growth factor (TGF)-ß1 were increased when BMSCs were co-cultured with CD8(+) T cells. The addition of specific inhibitors against PGE2 , IDO and TGF-ß partially restored the proliferation of CD8(+) T cells. Our results suggest that BMSCs suppress CD8(+) T cell-mediated activation by suppressing NKG2D expression and secretion of PGE2, IDO and TGF-ß. Our observations further confirm the feasibility of BMSCs as a potential adoptive cellular therapy in immune-mediated diseases such as graft-versus-host disease (GVHD).

Clinicopathological characteristics of vascular endothelial growth factor expression in uveal melanoma: A meta-analysis.

Angiogenesis is a potential prognostic factor that has been extensively investigated in patients with uveal melanoma (UM). Vascular endothelial growth factor (VEGF) expression is crucial in angiogenesis. However, there have been conflicting data regarding the clinicopathological data in UM. A meta-analysis was performed of all the germane literature to assess the clinicopathological characteristics of VEGF expression by combining separately estimated odds ratio (OR) values. Our combined results demonstrated that, according to the available studies, the expression of VEGF in UM was significantly higher compared to normal tissue [338 patients and 99 controls; OR=16.15, 95% confidence interval (CI): 8.65-30.12, P<0.00001]. When stratifying the studies by age (315 patients; OR=2.08, 95% CI: 1.19-3.62, P=0.01), cell type (423 patients; OR=0.54, 95% CI: 0.32-0.90, P=0.02), tumor size (222 patients; OR=0.30, 95% CI: 0.14-0.68, P=0.004) and scleral invasion (248 patients; OR=0.34, 95% CI: 0.15-0.78, P=0.01), significant clinicopathological information was provided. Our results indicated that VEGF expression in UM patients was significantly higher compared to that observed in controls. It was also significantly higher in patients who presented with scleral invasion and those who were aged <50 years. In addition, VEGF expression was higher in mixed-cell type and epithelioid-cell type UM and in patients with large-sized tumors.

A novel approach for enriching cancer stem cells from the human SW-13 adrenocortical carcinoma cell line.

The present study was undertaken to develop a new method for enriching cancer stem cells (CSCs) from the human adrenal cortical carcinoma (ACC) cell line SW-13. Given that the existence of CSCs in ACC causes resistance to conventional chemotherapies, treatment with cyclophosphamide was used for in vivo selection of CSCs in a BALB/c nude mouse tumor xenograft model established using the ACC cell line SW-13. The characteristics of CSCs in three generations of tumor xenografts were assessed for single-cell colony formation, flat colony formation, and cell sphere formation in serum-free suspension culture. The formation rates of single-cell colonies, flat colonies, and cell spheres were significantly higher for tumor xenograft cells treated with cyclophosphamide than for untreated engrafted tumor cells. Flow cytometry to examine expression of the CSC markers C-X-C chemokine receptor type-4 (CXCR4; CD184) and ATP-binding cassette sub-family G member-2 (ABCG2; CDw338) revealed markedly higher levels of CXCR4 and ABCG2 in cyclophosphamide-treated xenograft tumor cells compared to untreated tumor cells. Together, these results indicate that cyclophosphamide treatment of tumor xenograft cells caused enrichment of CSCs with a strong capability for self-renewal and proliferation. In this method, the administration of cyclophosphamide selectively kills cancer cells without toxicity to CSCs and thereby provides a practical approach for achieving the enrichment of CSCs in ACC.

Novel function of N,N-bis(2-chloroethyl)docos-13-enamide for reversal of multidrug resistance in tongue cancer.

Multidrug resistance (MDR) is a key element in the failure of chemotherapies, and development of agents to overcome MDR is crucial to improving cancer treatments. The overexpression of glutathione-S-transferases (GSTs) is one of the major mechanisms of MDR. Because some agents used in traditional Chinese medicine have strong antitumor effects coupled with low toxicity; we investigated the ability of N,N-bis(2-chloroethyl)docos-13-enamide (compound J), the synthesized analog of a highly unsaturated fatty acid from Isatis tinctoria L., to reverse the MDR induced by adriamycin (ADM) in TCA8113/ADM cells. We found that compound J significantly increased the cytotoxicity of ADM in TCA8113/ADM cells, with a reversal fold of 2.461. Analysis of the mechanisms through which compound J reversed MDR indicated that compound J significantly decreased the activity of GSTs and enhanced the depletion of GSH in TCA8113/ADM cells, but did not affect the P-glycoprotein (P-gp) efflux. Taken together, our data suggested that compound J was an excellent candidate for reversing MDR in cancer therapy.

Meta-analysis of the association of HLA-DRB1 with rheumatoid arthritis in Chinese populations.

BACKGROUND: Individual studies have reported different results regarding the association of HLA alleles with RA in Chinese populations. This study was performed to systematically summarize results on the association of HLA-DRB1 with rheumatoid arthritis (RA) in China. METHODS: We examined the case-control studies concerned about the relationship between HLA-DRB1 and RA and differences of clinical and laboratory parameters between the HLA-DR4 (DR4)+ and DR4- in RA patients in Chinese populations. Odds ratios (ORs) and weighted mean difference (WMD) with corresponding 95% confidence intervals (CI) was used to describe the relationship. RESULTS: 22 studies with 1690 cases and 1793 controls were included. Chinese populations with RA had significantly higher frequencies of HLA-DRB1*04, *0401, *0404, *0405 and *0410 than controls (ORDRB1*04 =4.19, 95% CI =3.44-5.11, p<0.00001; ORDRB1*0401 =2.53, 95% CI =1.54-4.16, p=0.0003; ORDRB1*0404 =2.28, 95% CI =1.28-4.06, p=0.005; ORDRB1*0405=3.71, 95% CI =2.52-5.45, p<0.00001; ORDRB1*0410 =2.99, 95% CI =1.25-7.14, p=0.01 respectively). As to laboratory parameters, Erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), Rheumatoid factor (RF), Anti-cyclic citrullinated peptide antibodies (Anti-CCP ) in patients with DR4+ were higher than patients with DR4- (WMD=0.26, 95% CI =0.15-0.37, p<0.00001; WMD = 0.26, 95% CI =0.12-0.41, p=0.0005; WMD = 0.44, 95% CI =0.23-0.65, p<0.00001; WMD = 0.58, 95% CI =0.24-0.91, p=0.0007 respectively). As to clinical features, there was no difference in duration of morning stiffness, number of swollen joints, number of joint tenderness, X-ray phases and joint function between the DR4+ and DR4- in RA patients. CONCLUSIONS: It was found that HLA-DRB1*04, *0401, *0404, *0405 and *0410 are risk factors for RA in Chinese populations. ESR, CRP, RF, Anti-CCP are different between the DR4+ and DR4- in RA patients in Chinese populations, while there's no difference for indexes of clinical features.

Real-time detection of single-living pancreatic beta-cell by laser tweezers Raman spectroscopy: high glucose stimulation.

Glucose acts as a beta-cell stimulus factor and leads to cellular responses that involve a large amount of biomolecule formation, relocation, and transformation. We hypothesize that information about these changes can be obtained in real-time by laser tweezers Raman spectroscopy. To test this hypothesis, repeated measurements designs in accordance with the application of Raman spectroscopy detection were used in the current experiment. Single rat beta-cells were measured by Raman spectroscopy in 2.8 mmol/l glucose culture medium as a basal condition. After stimulation with high glucose (20 mmol/l), the same cells were measured continuously. Each cell was monitored over a total time span of 25 min, in 5 min intervals. During this period of time, cells were maintained at an appropriate temperature controlled by an automatic heater, to provide near-physiological conditions. It was found that some significant spectral changes induced by glucose were taking place during the stimulation time course. The most noticeable changes were the increase of spectral intensity at the 1002, 1085, 1445, and 1655 cm(-1) peaks, mainly corresponding to protein and lipid. We speculate that these changes might have to do with beta-cell protein and lipid synthesis. Using laser tweezers Raman spectroscopy in combination with glucose stimulation, optical spectral information from rat beta-cells was received and analyzed.

Blumeaxanthene II, a novel xanthene from Blumea riparia DC.

A novel xanthene (1), blumeaxanthene II, was isolated from the aerial parts of Blumea riparia (Bl.) DC., a Chinese medicinal plant with hemostatic properties, and its structure was determined by extensive spectroscopic analyses. This compound is the first example of a halogenated xanthene from a natural plant, and a biosynthetic pathway was proposed. Blumeaxanthene II was also tested against Bel-7404 cells in vitro but was found to be only weakly cytotoxic.

Synergistic effects of Isatis tinctoria L. and tacrolimus in the prevention of acute heart rejection in mice.

Although immunosuppressive treatments are available for acute cardiac rejection no viable treatment exists for long-term cardiac graft failure. Moreover, the extended use of calcineurin inhibitor immunosuppressants, the mainstay of current treatment for cardiac transplantation, leads to significant side effects such as nephrotoxicity and an increased risk of cardiac disease. Because some agents used in Traditional Chinese Medicine (TCM) have strong immunosuppressive effects coupled with low toxicity, we investigated the effect of Compound K (K), the synthesized analogue of highly unsaturated fatty acids from Isatis tinctoria L., either as a single treatment or combined with tacrolimus (FK-506) on acute cardiac allograft rejection. We compared the ability of K alone, or in combination with FK-506, to inhibit acute heart transplant rejection both in vitro and in vivo. We found that the inhibition of lymphocyte proliferation was positively correlated with K concentration. K significantly reduced IL-2 and IFN-gamma expression levels and significantly inhibited lymphocyte proliferation in both a lymphocyte transformation test and a mixed lymphocyte reaction (MLR). We also found that the inhibitory effect of a combination of K and a sub-therapeutic dose of FK-506 (SubFK-506) was stronger than that of full-dose FK-506 alone. Oral administration of K reduced acute cardiac allograft rejection in mice and had no apparent toxicity. In vivo, the immunosuppressive effect of K combined with a half-dose of FK-506 was equivalent to that of a full-dose of FK-506 alone. K combined with a half-dose of FK-506 reduced the expression levels of IL-2 and IFN-gamma (both within the graft and in the recipients' serum) more effectively than a full-dose of FK-506. These results show that K has significant immunosuppressive effects both in vitro and in vivo. When used as a combination therapy with FK-506 we see a powerful inhibition of rejection with no obvious toxic side effects. The mechanism of action is postulated to involve the inhibition of IL-2 and IFN-gamma expressions by lymphocytes, rather than the activation of Tr1 cells via the production of IL-10.