A Comparative Study of the Bone Marrow- and Umbilical Cord-Derived Mesenchymal Stem Cells (MSCs) Efficiency on Generating MSC-Educated Macrophages (MEMs)

Background: Mesenchymal stem cells (MSCs) have gained much more attention in cell therapy and regenerative medicine due to their immunosuppressive effects. MSCs have interaction with other immune cells, such as macrophages (MQs). Bone marrow (BM)-derived MSCs can educate MQs toward MSC-educated MQs (MEMs) which possess an anti-inflammatory immunophenotype. Given this and based on the important limitations of BM collection, we hypothesized whether co-culture of MQs with umbilical cord (UC)-derived MSCs can result in the MEM phenotype. Methods: First, isolated monocytes cultured for five days to obtain M0 MQs. Then, they were co-cultured with either BM- or UC-MSCs under direct and indirect conditions. After three days of co-culture, MEM-specific surface markers, as well as the gene expression of inflammatory and anti-inflammatory cytokines, were evaluated. Results: Surface expression of CD163/CD206, as specific markers for M2 MQs, increased in MEMs after co-culture with BM- and UC-derived MSCs, while CD80/CD86 expression (specific markers for M1 MQs) didn’t change significantly. The mRNA expressions of PDL-1 as well as anti-inflammatory cytokines, including IL-6, IL-10, and TGFβ also increased in MEMs after co-culture of UC-MSCs compared to control MQs (p <.05), while the expression of IL-12 was significantly decreased (p<.001). Conclusions: To the best of our knowledge, this study shows for the first time that the co-culture of MQs with UC-derived MSCs efficiently contributes to the generation of MEMs even greater than BM-MSCs; shedding light on the promising potential of UC as an alternative source to educate MQs in vitro.     

Mild thermotolerance induced at 40 °C protects cells against hyperthermia-induced pro-apoptotic changes in Bcl-2 family proteins

Purpose: Despite clinical progress, mechanisms involved in cellular responses to low and high doses of hyperthermia are not entirely clear. This study investigates the role of Bcl-2 family proteins in control of the mitochondrial pathway of apoptosis during hyperthermia at 42–43?°C and the protective effect of a low dose adaptive survival response, mild thermotolerance induced at 40?°C.

Materials and methods: Levels of Bcl-2 family proteins were detected in HeLa cells by western blotting, caspase activation by spectrofluorimetry and apoptosis by chromatin condensation.

Results: Hyperthermia (42–43?°C) decreased total and mitochondrial expression of anti-apoptotic proteins Bcl-2 and Bcl-xL, while expression of pro-apoptotic proteins Bax, Bak, Puma and Noxa increased. Hyperthermia perturbed the equilibrium between these anti- and pro-apoptotic Bcl-2 family proteins in favour of pro-apoptotic conditions. Hyperthermia also caused activation of caspases-9 and -3, and chromatin condensation. Disruption of the balance between Bcl-2 family proteins was reversed in thermotolerant (40?°C) cells, thus favouring cell survival. Bcl-2/Bcl-xL inhibitor ABT-737 sensitised cells to apoptosis, which indicates that Bcl-2 family proteins play a role in hyperthermia-induced apoptosis. The adaptive response of mild thermotolerance (40?°C) was still able to protect cells against hyperthermia (42–43?°C) when Bcl-2/Bcl-xL were inhibited.

Conclusions: These results improve knowledge about the role of Bcl-2 family proteins in cellular apoptotic responses to hyperthermia (42–43?°C), as well as the adaptive survival response induced by exposure to mild stresses, such as a fever temperature (40?°C). This study could provide rationale to explore the manipulation of Bcl-2 family proteins for increasing tumour sensitivity to hyperthermia.     

Wnt5a‐Ror2 signaling in mesenchymal stem cells promotes proliferation of gastric cancer cells by activating CXCL16–CXCR6 axis

Wnt5a‐Ror2 signaling has been shown to play important roles in promoting aggressiveness of various cancer cells in a cell‐autonomous manner. However, little is known about its function in cancer‐associated stromal cells, including mesenchymal stem cells (MSCs). Thus, we examined the role of Wnt5a‐Ror2 signaling in bone marrow‐derived MSCs in regulating proliferation of undifferentiated gastric cancer cells. Coculture of a gastric cancer cell line, MKN45, with MSCs either directly or indirectly promotes proliferation of MKN45 cells, and suppressed expression of Ror2 in MSCs prior to coculture inhibits enhanced proliferation of MKN45 cells. In addition, conditioned media from MSCs, treated with control siRNA, but not siRNAs against Ror2, can enhance proliferation of MKN45 cells. Interestingly, it was found that expression of CXCL16 in MSCs is augmented by Wnt5a‐Ror2 signaling, and that recombinant chemokine (C‐X‐C motif) ligand (CXCL)16 protein can enhance proliferation of MKN45 cells in the absence of MSCs. In fact, suppressed expression of CXCL16 in MSCs or an addition of a neutralizing antibody against CXCL16 fails to promote proliferation of MKN45 cells in either direct or indirect coculture with MSCs. Importantly, we show that MKN45 cells express chemokine (C‐X‐C motif) receptor (CXCR)6, a receptor for CXCL16, and that suppressed expression of CXCR6 in MKN45 cells results in a failure of its enhanced proliferation in either direct or indirect coculture with MSCs. These findings indicate that Wnt5a‐Ror2 signaling enhances expression of CXCL16 in MSCs and, as a result, enhanced secretion of CXCL16 from MSCs might act on CXCR6 expressed on MKN45, leading to the promotion of its proliferation.     

Hyperthermia‐treated mesenchymal stem cells exert antitumor effects on human carcinoma cell line

BACKGROUND:

Mesenchymal stem cells (MSCs) possess the potential for differentiation into multilineages. MSCs have been reported to play a role as precursors for tumor stroma in providing a favorable environment for tumor progression. Hyperthermia destroys cancer cells by raising the temperature of tumor‐loaded tissue to 40°C to 43°C and causes indirect sensitizing effects when combined with chemo‐ and/or radiotherapy. However, how hyperthermia affects the tumor‐supportive stroma is unknown. Here, the authors investigated the effects of hyperthermia‐treated MSCs, from different sources, on the human ovarian cancer cell line SK‐OV‐3.

METHODS:

MSCs from adipose tissue and amniotic fluid were untreated or heat‐treated (HS‐MSCs). The culture supernatant of each treatment group was collected and transferred to the SK‐OV‐3 cells.

RESULTS:

The morphological analysis and cell proliferation assay showed a reduced viability of the tumor cells in the conditioned medium with the HS‐MSCs. Further investigations revealed that the conditioned medium of the HS‐MSCs induced a higher nuclear condensation and a greater number of sub‐G1 cells among the tumor cells. Analysis of the mRNA expression demonstrated that the conditioned medium of the HS‐MSCs induced up‐regulation or down‐regulation of several tumor‐associated molecules. Finally, the cytokine array of each conditioned medium showed that angiogenin, insulin‐like growth factor binding protein 4, neurotrophin 3, and chemokine (C‐C motif) ligand 18 are involved as main factors.

CONCLUSIONS:

This study showed that the conditioned medium of the HS‐MSCs exerted a suppressive effect on tumor progression and malignancy, suggesting that hyperthermia enables tumor stromal cells to provide a sensitizing environment for tumor cells to undergo cell death. Cancer 2009. © 2009 American Cancer Society.     

Deep hyperthermia with the HYPERcollar system combined with irradiation for advanced head and neck carcinoma – a feasibility study

Purpose: Radiotherapy (RT) treatment of locally-advanced and recurrent head and neck carcinoma (HNC) results in disappointing outcomes. Combination of RT with cisplatin or cetuximab improves survival but the increased toxicity and patient's comorbidity warrant the need for a less-toxic radiosensitizer. Stimulated by several randomized studies demonstrating the radio-sensitizing effect of hyperthermia, we developed the HYPERcollar. Here, we report early experience and toxicity in patients with advanced HNC.

Methods and materials: 119 hyperthermia treatments given to 27 patients were analyzed. Hyperthermia was applied once a week by the HYPERcollar aimed at achieving 39–43?°C in the target area, up to patients’ tolerance. Pre-treatment planning was used to optimize treatment settings. When possible, invasive thermometry catheters were placed.

Results: Mean power applied during the 119 hyperthermia treatments ranged from 120 to 1007?W (median 543?W). 15 (13%) hyperthermia treatments were not fully completed due to: pain allocated to hyperthermia (6/15), dyspnea from sticky saliva associated with irradiation (2/15) and unknown reasons (7/15). No severe complications or enhanced thermal or mucosal toxicities were observed. Excluding post-operative treatment, response rates after 3?months were 46% (complete) and 7% (partial).

Conclusion: Hyperthermia with the HYPERcollar proved to be safe and feasible with good compliance and promising outcome.     

Exposure to febrile-range hyperthermia potentiates Wnt signalling and epithelial–mesenchymal transition gene expression in lung epithelium

Background: As environmental and body temperatures vary, lung epithelial cells experience temperatures significantly different from normal core temperature. Our previous studies in human lung epithelium showed that: (i) heat shock accelerates wound healing and activates profibrotic gene expression through heat shock factor-1 (HSF1); (ii) HSF1 is activated at febrile temperatures (38–41?°C) and (iii) hypothermia (32?°C) activates and hyperthermia (39.5?°C) reduces expression of a subset of miRNAs that target protein kinase-Cα (PKCα) and enhance proliferation.

Methods: We analysed the effect of hypo- and hyperthermia exposure on Wnt signalling by exposing human small airway epithelial cells (SAECs) and HEK293T cells to 32, 37 or 39.5?°C for 24?h, then analysing Wnt-3a-induced epithelial–mesenchymal transition (EMT) gene expression by qRT-PCR and TOPFlash reporter plasmid activity. Effects of miRNA mimics and inhibitors and the HSF1 inhibitor, KNK437, were evaluated.

Results: Exposure to 39.5?°C for 24?h increased subsequent Wnt-3a-induced EMT gene expression in SAECs and Wnt-3a-induced TOPFlash activity in HEK293T cells. Increased Wnt responsiveness was associated with HSF1 activation and blocked by KNK437. Overexpressing temperature-responsive miRNA mimics reduced Wnt responsiveness in 39.5?°C-exposed HEK293T cells, but inhibitors of the same miRNAs failed to restore Wnt responsiveness in 32?°C-exposed HEK293T cells.

Conclusions: Wnt responsiveness, including expression of genes associated with EMT, increases after exposure to febrile-range temperature through an HSF1-dependent mechanism that is independent of previously identified temperature-dependent miRNAs. This process may be relevant to febrile fibrosing lung diseases, including the fibroproliferative phase of acute respiratory distress syndrome (ARDS) and exacerbations of idiopathic pulmonary fibrosis (IPF).     

Hyperthermia on mesenchymal stem cells (MSCs) can sensitize tumor cells to undergo cell death†

Hyperthermia, the procedure of raising the temperature of tumor-loaded tissue to 40°–43°C, has been applied to various established cancer treatments. Although the mechanism of hyperthermia in cancer treatment is well-known, there are few or no studies regarding the effect of hyperthermia on the tumor-supportive stroma. Mesenchymal stem cells (MSCs) display the potential for differentiation into various tissues. MSCs are also reported to play a role as potential precursors for tumor stroma in providing a favorable environment for tumor progression. Here, we investigated the effects of hyperthermia-treated MSCs on the viability and growth of cancer cells. Culture supernatants from non-shocked or heat-shocked MSCs (NS-MSCs or HS-MSCs) were added to MCF7 cells. Morphological analysis and cell proliferation assay showed the reduced viability and growth of MCF7 cells by addition of culture medium conditioned by HS-MSCs. Additionally, exposure to the conditioned medium by HS-MSCs induced cell cycle arrest at G2/M phase, increased MHC class I, Fas receptor, and TNF-R expressions, and decreased MDR1 expression in the MCF7 cells. In particular, the conditioned medium of HS-MSCs accelerated the inhibition of tumor cell growth by several chemotherapeutic drugs. These data present new aspects of hyperthermia in cancer treatment, suggesting that hyperthermia can enable tumor stroma provide a sensitizing environment for tumor cells to undergo cell death.     

Cross-talk between chronic lymphocytic leukemia (CLL) tumor B cells and mesenchymal stromal cells (MSCs): implications for neoplastic cell survival

Leukemic cells from Chronic Lymphocytic Leukemia (CLL) patients interact with stromal cells of the surrounding microenvironment. Mesenchymal Stromal Cells (MSCs) represent the main population in CLL marrow stroma, which may play a key role for disease support and progression. In this study we evaluated whether MSCs influence in vitro CLL cell survival. MSCs were isolated from the bone marrow of 46 CLL patients and were characterized by flow cytometry analysis. Following co-culture of MSCs and leukemic B cells, we demonstrated that MSCs were able to improve leukemic B cell viability, this latter being differently dependent from the signals coming from MSCs. In addition, we found that the co-culture of MSCs with leukemic B cells induced an increased production of IL-8, CCL4, CCL11, and CXCL10 chemokines.As far as drug resistance is concerned, MSCs counteract the cytotoxic effect of Fludarabine/Cyclophosphamide administration in vivo, whereas they do not protect CLL cells from the apoptosis induced by the kinase inhibitors Bafetinib and Ibrutinib. The evidence that leukemic clones are conditioned by environmental stimuli suggest new putative targets for therapy in CLL patients.     

Inhibitory effect and mechanism of mesenchymal stem cells on melanoma cells

Purpose

To explore the inhibitory effect and mechanism of MSCs on melanoma proliferation.

Methods

The inhibitory effect of MSCs on melanoma A375 cells was detected by co-culture and conditioned medium (CM) experiments using MTT method. The cell cycle was analyzed by flow cytometry. Then, Western Blot experiment detected the expression of proteins related to NF-κB signaling in A375 cells. The expression of IL-1Ra in MSCs was proved by RT-PCR. The over-expression and silencing vector pcDNA3.1-EGFP-IL-1Ra and pGPH1-IL-1R were constructed and transfected into MSCs cells. After that, the changes of inhibitory effect and cell cycle from MSCs-S and MSCs-O CM on A375 cells were explored. The expression of proteins related to NF-κB signaling in A375 cells after MSCs-S or MSCs-O CM treatment was detected by Western Blot. MSCs, MSCs-S, or MSCs-O and A375 cells were co-injected into nude mice under the arms, the growth of tumor was observed, the frozen sections were made, and H&E staining of tumor tissue was performed.

Results

The proliferation of A375 cells was inhibited and the cell cycle of A375 was arrested by MSCs. The expressions of cytokines related to NF-κB signaling were down-regulated. Over-expression and silence of Interleukin 1 receptor antagonist (IL-1Ra), specifically blocking activation of NF-κB signaling, indicated that inhibitory effect from MSCs was enhanced or weakened respectively, which suggested that IL-1Ra was involved in the inhibitory effect. In vivo, tumor initiation and growth were significantly inhibited when A375 cells were co-injected with MSCs into nude mice, which were related to the expression level of IL-1Ra.

Conclusion

MSCs could inhibit the proliferation and tumor initiation of melanoma A375 cells through NF-κB signaling. MSCs could secret IL-1Ra and inhibit expressions of NF-κB signaling-related factors of tumor cells, and cause cell cycle arrest in G1 phase.

     

Bone marrow-derived mesenchymal stem cell-secreted IL-8 promotes the angiogenesis and growth of colorectal cancer

Mesenchymal stem cells (MSCs) have recently been shown to home to tumors and contribute to the formation of the tumor-associated stroma. In addition, MSCs can secrete paracrine factors to facilitate tumor progression. However, the involvement of MSC-derived cytokines in colorectal cancer (CRC) angiogenesis and growth has not been clearly addressed. In this study, we report that interleukin-8 (IL-8) was the most highly upregulated pro-angiogenic factor in MSCs co-cultured with CRC cells and was expressed at substantially higher levels in MSCs than CRC cells. To evaluate the effect of MSC-derived IL-8 on CRC angiogenesis and growth, we used MSCs that expressed small hairpin (interfering) RNAs (shRNA) targeting IL-8 (shIL-8-MSCs). We found that MSC-secreted IL-8 promoted human umbilical vein endothelial cell (HUVEC) proliferation and migration, tube-formation ability and CRC cell proliferation. Additionally, in vivo studies showed that MSCs promoted tumor angiogenesis partially through IL-8. Taken together, these findings suggest that IL-8 secreted by MSCs promotes CRC angiogenesis and growth and can therefore serve as a potential novel therapeutic target.     

Localised hyperthermia in rodent models using an MRI-compatible high-intensity focused ultrasound system

Abstract

Purpose: Localised hyperthermia in rodent studies is challenging due to the small target size. This study describes the development and characterisation of an MRI-compatible high-intensity focused ultrasound (HIFU) system to perform localised mild hyperthermia treatments in rodent models. Material and methods: The hyperthermia platform consisted of an MRI-compatible small animal HIFU system, focused transducers with sector-vortex lenses, a custom-made receive coil, and means to maintain systemic temperatures of rodents. The system was integrated into a 3T MR imager. Control software was developed to acquire images, process temperature maps, and adjust output power using a proportional-integral-derivative feedback control algorithm. Hyperthermia exposures were performed in tissue-mimicking phantoms and in a rodent model (n?=?9). During heating, an ROI was assigned in the heated region for temperature control and the target temperature was 42?°C; 30?min mild hyperthermia treatment followed by a 10-min cooling procedure was performed on each animal. Results: 3D-printed sector-vortex lenses were successful at creating annular focal regions which enables customisation of the heating volume. Localised mild hyperthermia performed in rats produced a mean ROI temperature of 42.1?±?0.3?°C. The T10 and T90 percentiles were 43.2?±?0.4?°C and 41.0?±?0.3?°C, respectively. For a 30-min treatment, the mean time duration between 41–45?°C was 31.1?min within the ROI. Conclusions: The MRI-compatible HIFU system was successfully adapted to perform localised mild hyperthermia treatment in rodent models. A target temperature of 42?°C was well-maintained in a rat thigh model for 30?min.     

Analysis of clinical data to determine the minimum number of sensors required for adequate skin temperature monitoring of superficial hyperthermia treatments

Purpose: Tumor response and treatment toxicity are related to minimum and maximum tissue temperatures during hyperthermia, respectively. Using a large set of clinical data, we analyzed the number of sensors required to adequately monitor skin temperature during superficial hyperthermia treatment of breast cancer patients.

Methods: Hyperthermia treatments monitored with >60 stationary temperature sensors were selected from a database of patients with recurrent breast cancer treated with re-irradiation (23?×?2?Gy) and hyperthermia using single 434?MHz applicators (effective field size 351–396?cm²). Reduced temperature monitoring schemes involved randomly selected subsets of stationary skin sensors, and another subset simulating continuous thermal mapping of the skin. Temperature differences (ΔT) between subsets and complete sets of sensors were evaluated in terms of overall minimum (T_min) and maximum (T_max) temperature, as well as T90 and T10.

Results: Eighty patients were included yielding a total of 400 hyperthermia sessions. Median ΔT was?<0.01?°C for T90, its 95% confidence interval (95%CI) decreased to ≤0.5?°C when?>50 sensors were used. Subsets of?<10 sensors result in underestimation of T_max up to ?2.1?°C (ΔT 95%CI), which decreased to ?0.5?°C when?>50 sensors were used. Thermal profiles (8–21 probes) yielded a median ΔT?T_max, with a 95%CI of ?0.2?°C and 0.4?°C, respectively. The detection rate of T_max?≥43?°C is?≥85% while using?>50 stationary sensors or thermal profiles.

Conclusions: Adequate coverage of the skin temperature distribution during superficial hyperthermia treatment requires the use of?>50 stationary sensors per 400?cm² applicator. Thermal mapping is a valid alternative.     

Combined treatment of IL-1α and TNF-α potentiates the antitumour effect of hyperthermia

Vasoactive cytokines, such as IL-1α and TNF-α, modulate the homeostatic state at the endothelial surface and cause various types of pathological damage in vascular systems. We investigated the potential therapeutic effects of IL-1α and TNF-α in combination with hyperthermia on SCK tumours grown in the legs of A/J mice. We first determined the effect of cytokines on tumour blood perfusion with the ⁸⁶Rb uptake method. When the host mice were given an i.p. injection of 25 μg/kg IL-1α or 50 μg/kg TNF-α, the tumour blood perfusion markedly declined to 46 and 82% of control, respectively. The combination of IL-1α and TNF-α reduced the ⁸⁶Rb uptake to 41% of control. Hyperthermia at 42·5°C for 1 h reduced the tumour blood flow to 71% of control. The tumour blood perfusion decreased further to 20% of control when the tumours were heated for 1 h at 42.5°C starting 4h after the injection of both IL-1α and TNF-α. The changes in clonogenic cell numbers in SCK tumours, as determined by the in vivo-in vitro assay, following various treatments was also investigated. At 4h after an i.p. injection of 25 μg/kg IL-1α or 50 μg/kg TNF-α, the clonogenicity of SCK tumours significantly decreased to 29 or 37% of control, respectively. Heating at 42.5°C for 1h caused a decline in the clonogenic cell number to 30% of control. When both IL-1α and TNF-α were given and tumours were heated 4h later at 42·5°C for 1h, the clonogenic cell number markedly declined to 0.4% of control. The time needed for control tumours to reach 4x their initial volume was about 3 days, and treatment with IL-1α or hyperthermia alone induced a tumour delay growth by about 1 day. The combined injection of IL-1α and TNF-α followed by a heating at 42·5°C for 1h delayed the tumour growth by 6 days. The results in this study suggest that prior impairment of blood circulation by the combined treatment of IL-1α and TNF-α potentiates hyperthermic damage in tumours.     

Differential expression of mRNA in human monocytes following interaction with human colon cancer cells

Monocytes are known to differentiate into tissue-specific macrophages in response to the tissue environment, and it has been suggested that tumor-associated macrophages might promote angiogenesis. Therefore, the factors associated with monocyte differentiation into tumor-associated macrophages may become new targets for cancer therapy. However, these factors remain unclear in human colon cancer. The aim of this study was to identify the factors associated with human monocyte differentiation into tumor-associated macrophages at human colon cancer sites. MATERIALS AND METHODS: A human monocyte cell line (THP-1) was co-cultured with a human colon cancer cell line (DLD-1) and mRNA expression was analyzed by quantitative real-time PCR. RESULTS: In THP-1 cells, monocyte chemotactic protein (MCP)-1 mRNA expression increased in a time-dependent manner from day 3 after co-culture with DLD-1 cells; furthermore, expression of vascular endothelial growth factor (VEGF)-A, tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-8 mRNA was increased from day 5. This increase in mRNA expression in the THP-1 cells was attributable to the presence of the DLD-1 cells. Therefore, MCP-1, VEGF-A, TNF-α, IL-1β, and IL-8 are suggested to be associated with differentiation of human monocytes into tumor-associated macrophages at human colon cancer sites.     

高温对舌癌细胞体外增殖活性的影响

 目的 研究高温对舌癌细胞株增殖活性的影响，探讨高温治癌的作用机制。方法 对舌癌细胞株Tca-8113进行体外43℃加温，采用流式细胞术及MTT法研究加温后舌癌细胞增殖活性的变化。结果 G0～G1，S期细胞百分数在加温后无明显改变，而加温10分钟G2～M期细胞百分数则迅速降低，但它不随加温时间的延长而改变。结论 加温能使分裂期细胞减少，细胞的增殖活性降低，从而抑制细胞的增殖，达到治疗肿瘤的目的。     

小胶质细胞及其细胞因子对乳腺癌细胞的影响

目的:体外培养小胶质N9细胞并观察其条件培养液(conditioned medium,CM)对乳腺癌MCF-7细胞生存率的影响,以探讨小胶质细胞在脑转移癌中的作用机制.方法:混合培养N9细胞和MCF-7细胞,在倒置光学显微镜下观察细胞形态学的变化.采用脂多糖(lipopolysaccharide,LPS)、MCF-7细胞及其条件培养液MCF-7-CM刺激N9细胞获得N9-CM; ELISA法检测N9-CM中肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-17 (interleukin-17,IL-17)的表达,Griess体系检测一氧化氮(nitric oxide,NO)的生成量.将N9-CM按不同浓度(10％、25％、50％、75％和100％)作用于MCF-7细胞,MTT法检测Ng-CM对MCF-7细胞生存率的影响.结果:MCF-7细胞与N9细胞共培养,N9细胞由静止态变为激活态;LPS、MCF-7细胞及其上清液均可以诱导N9细胞产生TNF-α、IL-17和NO;N9-CM浓度达50％～100％时对MCF-7细胞生长有抑制作用,且作用强度呈时间-浓度相关性.结论:小胶质细胞可以被转移到CNS的乳腺癌细胞激活并发挥肿瘤抑制作用.其作用强度与小胶质细胞分泌NO及细胞因子TNF-α和IL-17的浓度和作用时间呈正相关性.     

胃癌细胞外泌体调控M2型巨噬细胞极化及对胃癌细胞增殖和迁移的影响

目的:探究胃癌细胞外泌体调控M2型巨噬细胞极化及对胃癌细胞增殖和迁移能力的影响。方法:差速离心法提取胃黏膜上皮细胞GES-1和胃癌细胞株MGC-803、SGC-7901外泌体,透射电镜和Western blot鉴定外泌体,将U937细胞与PMA共孵育诱导U937细胞分化为巨噬细胞的同时分别与PBS、GES-1 exo、MGC-803 exo、SGC-7901 exo、IL-4孵育48 h,流式细胞术检测孵育后各组细胞CD206和HLA-DR表达,qRT-PCR检测CCL17、CXCL8、IL-10、TGF-β、TNF-α、IL-6、IL-1β表达,Western blot检测p-NF-κB、NF-κB、IκBα表达水平,将各组诱导后的U937细胞与MGC-803、SGC-7901共培养后,收集MGC-803、SGC-7901细胞,MTT检测各组细胞的增殖能力,Transwell小室法检测各组细胞的迁移能力。结果:差速离心法提取的外泌体符合外泌体的形态特征,并且在激光共聚焦显微镜下观察到外泌体可被U937细胞摄取,与MGC-803 exo、SGC-7901 exo共孵育后的U937细胞CD206显著高表达,HLA-DR低表达（P＜0.05）,CCL17、CXCL8、IL-10、TGF-β表达显著上调（P＜0.05）,TNF-α、IL-6、IL-1β表达显著下调（P＜0.05）,NF-κB磷酸化水平显著增加（P＜0.05）,IκBα表达显著增加（P＜0.05）,并且与MGC-803 exo、SGC-7901 exo诱导极化后的巨噬细胞共培养组MGC-803、SGC-7901细胞增殖和迁移能力显著增强（P＜0.05）。结论:胃癌细胞能够通过激活NF-κB信号通路促进M2型巨噬细胞极化进而诱导MGC-803、SGC-7901细胞增殖和迁移能力的增强。     

Heme oxygenase-1 (Hsp32) is involved in the protection of small intestine by whole body mild hyperthermia from ischemia/reperfusion injury in rat

Aim: The aim of the present study was to explore whether heme oxygenase-1 (HO-1) is involved in the hyperthermia-provided protection of the small intestine from ischemia/reperfusion injury in rats.

Methods: Intestinal damage was induced in male Sprague-Dawley rats by clamping both the superior mesenteric artery and the celiac trunk for 30?min, followed by reperfusion. Whole-body hyperthermia was induced in anesthetized rats by placement in a temperature-controlled water bath. Whole-body hyperthermia to a core temperature of 42–43°C for 15?min was followed by passive cooling. We started the hyperthermic treatment 6?h before the vascular clamping. The severity of the mucosal injury was evaluated by several biochemical markers and histological findings. Hyperthermia-induced heat-shock proteins were detected by Western blotting. We also investigated the effect of zinc protoporphyrin IX (an HO-1 inhibitor) on the protective effect of hyperthermia.

Results: The rats, which were killed after ischemia/reperfusion, had severe intestinal inflammation. Hyperthermia significantly induced the production of Hsp70 and HO-1 in intestinal mucosa and significantly reduced ischemia/reperfusion-induced mucosal injury. The combination of zinc protoporphyrin IX with hyperthermia extinguished the protective effects of hyperthermia on ischemia/reperfusion injury.

Conclusion: Hyperthermia protects against ischemia/reperfusion injury in rat small intestine through the expression of heat-shock proteins, especially HO-1.