Zongbing You

The number and microlocalization of tumor-associated immune cells are associated with patient's survival time in non-small cell lung cancer

BackgroundTumor microenvironment is composed of tumor cells, fibroblasts, endothelial cells, and infiltrating immune cells. Tumor-associated immune cells may inhibit or promote tumor growth and progression. This study was conducted to determine whether the number and microlocalization of macrophages, mature dendritic cells and cytotoxic T cells in non-small cell lung cancer are associated with patients survival time.MethodsNinety-nine patients with non-small cell lung cancer (NSCLC) were included in this retrospective study. Paraffin-embedded NSCLC specimens and their clinicopathological data including up to 8-year follow-up information were used. Immunohistochemical staining for CD68 (marker for macrophages), CD83 (marker for mature dendritic cells), and CD8 (marker for cytotoxic T cells) was performed and evaluated in a blinded fashion. The numbers of immune cells in tumor islets and stroma, tumor islets, or tumor stroma were counted under a microscope. Correlation of the cell numbers and patients survival time was analyzed using the Statistical Package for the Social Sciences (version 13.0).ResultsThe numbers of macrophages, mature dendritic cells and cytotoxic T cells were significantly more in the tumor stroma than in the tumor islets. The number of macrophages in the tumor islets was positively associated with patients survival time, whereas the number of macrophages in the tumor stroma was negatively associated with patients survival time in both univariate and multivariate analyses. The number of mature dendritic cells in the tumor islets and stroma, tumor islets only, or tumor stroma only was positively associated with patients survival time in a univariate analysis but not in a multivariate analysis. The number of cytotoxic T cells in the tumor islets and stroma was positively associated with patients survival time in a univariate analysis but not in a multivariate analysis. The number of cytotoxic T cells in the tumor islets only or stroma only was not associated with patients survival time.ConclusionsThe number of macrophages in the tumor islets or stroma is an independent predictor of survival time in NSCLC patients. Counting macrophages in the tumor islets or stroma is more useful in predicting patients survival time than counting mature dendritic cells or cytotoxic T cells.

Bone Morphogenetic Protein (BMP)-6 Signaling and BMP Antagonist Noggin in Prostate Cancer

It has been proposed that the osteoblastic nature of prostate cancer skeletal metastases is due in part to elevated activity of bone morphogenetic proteins (BMPs). BMPs are osteoinductive morphogens, and elevated expression of BMP-6 correlates with skeletal metastases of prostate cancer. In this study, we investigated the expression levels of BMPs and their modulators in prostate, using microarray analysis of cell cultures and gene expression. Addition of exogenous BMP-6 to DU-145 prostate cancer cell cultures inhibited their growth by up-regulation of several cyclin-dependent kinase inhibitors such as p21/CIP, p18, and p19. Expression of noggin, a BMP antagonist, was significantly up-regulated by BMP-6 by microarray analysis and was confirmed by quantitative reverse transcription-polymerase chain reaction and at the protein level. Noggin protein was present in prostate biopsies and localized to the epithelial components of prostate by immunohistochemistry. Recombinant noggin inhibited the function of BMP-6, suggesting a negative feedback regulation of BMP activity and indicating a strategy for the development of a novel therapeutic target in the treatment of painful osteosclerotic bone metastases of prostate cancer.

Interleukin-17 and Prostaglandin E2 Are Involved in Formation of an M2 Macrophage-Dominant Microenvironment in Lung Cancer

Introduction: Tumor-associated macrophages (TAMs) are divided into M1 and M2 macrophages. M1 macrophages inhibit tumor growth, whereas M2 macrophages promote tumor growth and metastasis. The aim of this study was to examine the possible causes leading to the formation of an M2-macrophage–dominant tumor microenvironment in non–small-cell lung cancer. Methods: Forty-eight archived lung tumor samples were examined for the expression of interleukin-17 (IL-17) receptors, IL-17 receptor A (IL-17RA) and IL-17 receptor C (IL-17RC), and the number of TAMs using immunohistochemical staining. Twenty fresh lung tumors and matched normal lung tissues were examined for expression of IL-17, cyclooxygenase-2, and prostaglandin E2 (PGE2), using enzyme-linked immunosorbent assay and Western blot analysis. Macrophage-migration assays were performed using fresh lung tumor tissues and IL-17 as chemoattractants. Induction of M2-macrophage differentiation was analyzed using real-time quantitative polymerase chain reaction. Results: TAMs expressed IL-17RA and IL-17RC. Lung tumors expressed higher levels of IL-17, cyclooxygenase-2, and PGE2, compared with normal lung tissues. Lung tumor tissues attracted migration of mouse RAW264.7 macrophages and primary peritoneal macrophages through IL-17, which was mediated by IL-17RA and IL-17RC. IL-17 did not induce either M1- or M2-macrophage differentiation. However, human lung cancer A549 cells strongly induced M2-macrophage differentiation of RAW264.7 macrophages when the two cell lines were cocultured. The inductive factor secreted by A549 cells was identified to be PGE2. Conclusions: IL-17 recruits macrophages, and PGE2 induces M2-macrophage differentiation, hence the increased levels of IL-17 and PGE2 in lung cancer contribute to the formation of an M2-macrophage–dominant tumor microenvironment.

Expression of interleukin-17RC protein in normal human tissues

Background Interleukin-17 (IL-17) cytokines and receptors play an important role in many autoimmune and inflammatory diseases. IL-17 receptors IL-17RA and IL-17RC have been found to form a heterodimer for mediating the signals of IL-17A and IL-17F cytokines. While the function and signaling pathway of IL-17RA has been revealed, IL-17RC has not been well characterized. The function and signaling pathway of IL-17RC remain largely unknown. The purpose of the present study was to systematically examine IL-17RC protein expression in 53 human tissues. Results IL-17RC expression in 51 normal human tissues and two benign tumors (i.e., lymphangioma and parathyroid adenoma) on the tissue microarrays was determined by immunohistochemical staining, using two polyclonal antibodies against IL-17RC. IL-17RC protein was expressed in many cell types including the myocardial cells, vascular and lymphatic endothelial cells, glandular cells (of the adrenal, parathyroid, pituitary, thyroid, pancreas, parotid salivary, and subepidermal glands), epithelial cells (of the esophagus, stomach, intestine, anus, renal tubule, breast, cervix, Fallopian tube, epididymis, seminal vesicle, prostate, gallbladder, bronchus, lung, and skin), oocytes in the ovary, Sertoli cells in the testis, motor neurons in the spinal cord, autonomic ganglia and nerves in the intestine, skeletal muscle cells, adipocytes, articular chondrocytes, and synovial cells. High levels of IL-17RC protein expression were observed in most vascular and lymphatic endothelium and squamous epithelium. The epithelium of the breast, cervix, Fallopian tube, kidney, bladder and bronchus also expressed high levels of IL-17RC, so did the glandular cells in the adrenal cortex, parotid salivary and subepidermal glands. In contrast, IL-17RC protein was not detectable in the smooth muscle cells, fibroblasts, antral mucosa of the stomach, mucosa of the colon, endometrium of the uterus, neurons of the brain, hepatocytes, or lymphocytes. Nevertheless, IL-17RC protein was expressed in the vascular endothelium within the tissues where the IL-17RC-negative cells resided. Conclusion IL-17RC protein is expressed in most human tissues, the function of which warrants further investigation.

Is Chemical Incompatibility Responsible for Chondrocyte Death Induced by Local Anesthetics

Background Chondrolysis associated with intra-articular administration of local anesthetics has been attributed to chondrocyte death induced by the local anesthetics. The mechanism of how the local anesthetics cause chondrocyte death is not clear. Purpose This study was conducted to determine whether and how the local anesthetics cause chondrocyte death. Study Design Controlled laboratory study. Methods Bovine articular chondrocytes in suspension culture were treated for 1 hour with phosphate-buffered saline or phosphate-buffered saline/medium mixture (as controls); 1% lidocaine alone; 0.25% to 0.5% bupivacaine alone; phosphate-buffered saline with pH values of 4.5, 3.8, 3.4, and 2.4; or mixtures of the local anesthetics and cell culture medium or human synovial fluid. Chondrocyte viability was analyzed by flow cytometry using the LIVE/DEAD Viability/Cytotoxicity Kit. Results In 1% lidocaine-alone or 0.25% to 0.5% bupivacaine-alone groups, the rate of cell death was 11.8% to 13.3% of bovine articular chondrocytes, whereas the phosphate-buffered saline control had 8.4% of cell death. Increased chondrocyte death was only found when the pH value of phosphate-buffered saline dropped to ≤3.4. In contrast, when bupivacaine was mixed with cell culture medium, needle-like crystals were formed, which was accompanied with 100% death of chondrocytes. Lidocaine did not form visible crystals when it was mixed with culture medium, but the mixtures caused death of over 96% of chondrocytes (P < .001). Conclusion Less than 5% of chondrocyte death was attributable to the anesthetics when applied to the cells alone or in phosphate-buffered saline—diluted solution. Acidity (as low as pH 3.8) or epinephrine in the anesthetic solutions could not account for chondrocyte death. However, chemical incompatibility between the local anesthetics and cell culture medium or human synovial fluid may be the cause of chondrocyte death. Clinical Relevance Intra-articular administration of lidocaine and bupivacaine is not an indicated usage of either anesthetic, although such a usage has become a common practice. Physicians should be aware of the potential incompatibility of the drug and synovial fluid.

Interleukin-17 Promotes Formation and Growth of Prostate Adenocarcinoma in Mouse Models

The contributions of interleukin (IL)-17 to cancer remain unclear and somewhat controversial. We took a genetic approach to explore its role in prostate cancers by interbreeding IL-17 receptor C (IL-17RC)-deficient mice with mice that are conditionally mutant for PTEN, one established preclinical model for prostate cancer. Mice that were IL-17RC-deficient (IL-17RC(-)) displayed prostates that were smaller than mice that maintained IL-17RC expression (IL-17RC(+)). In addition, IL-17RC(-) mice developed a reduced number of invasive prostate adenocarcinomas with lower rates of cellular proliferation and higher apoptosis than IL-17RC(+) mice. Moreover, the fibromuscular stroma surrounding prostatic glands was relatively thicker in IL-17RC(-) mice and was associated with decreased matrix metalloproteinase (Mmp)7 expression and increased Timp1, 2, and 4 expression, whereas administration of recombinant mouse IL-17 induced prostatic expression of Mmp7. Taken together, our results suggested that IL-17 promotes the formation and growth of prostate adenocarcinoma, and that an IL-17-MMP7 signaling axis is required for the transition of prostatic intraepithelial neoplasia to frank adenocarcinoma.

Midkine is a NF-κB-inducible gene that supports prostate cancer cell survival

BackgroundMidkine is a heparin-binding growth factor that is over-expressed in various human cancers and plays important roles in cell transformation, growth, survival, migration, and angiogenesis. However, little is known about the upstream factors and signaling mechanisms that regulate midkine gene expression.MethodsTwo prostate cancer cell lines LNCaP and PC3 were studied for their expression of midkine. Induction of midkine expression in LNCaP cells by serum, growth factors and cytokines was determined by Western blot analysis and/or real-time quantitative reverse-transcription – polymerase chain reaction (RT-PCR). The cell viability was determined by the trypan blue exclusion assay when the LNCaP cells were treated with tumor necrosis factor alpha (TNFα) and/or recombinant midkine. When the LNCaP cells were treated with recombinant midkine, activation of intracellular signalling pathways was determined by Western blot analysis. Prostate tissue microarray slides containing 129 cases (18 normal prostate tissues, 40 early stage cancers, and 71 late stage cancers) were assessed for midkine expression by immunohistochemical staining.ResultsWe identified that fetal bovine serum, some growth factors (epidermal growth factor, androgen, insulin-like growth factor-I, and hepatocyte growth factor) and cytokines (TNFα and interleukin-1beta) induced midkine expression in a human prostate cancer cell line LNCaP cells. TNFα also induced midkine expression in PC3 cells. TNFα was the strongest inducer of midkine expression via nuclear factor-kappa B pathway. Midkine partially inhibited TNFα-induced apoptosis in LNCaP cells. Knockdown of endogenous midkine expression by small interfering RNA enhanced TNFα-induced apoptosis in LNCaP cells. Midkine activated extracellular signal-regulated kinase 1/2 and p38 mitogen-activated protein kinase pathways in LNCaP cells. Furthermore, midkine expression was significantly increased in late stage prostate cancer, which coincides with previously reported high serum levels of TNFα in advanced prostate cancer.ConclusionThese findings provide the first demonstration that midkine expression is induced by certain growth factors and cytokines, particularly TNFα, which offers new insight into understanding how midkine expression is increased in the late stage prostate cancer.

Liposomal extended-release bupivacaine for postsurgical analgesia.

When physicians consider which analgesia to use postsurgery, the primary goal is to relieve pain with minimal adverse side effects. Bupivacaine, a commonly used analgesic, has been formulated into an aqueous suspension of multivesicular liposomes that provide long-lasting analgesia for up to 72 hours, while avoiding the adverse side effects of opioids. The increased efficacy of liposomal extended-release bupivacaine, compared to bupivacaine hydrochloride, has promoted its usage in a variety of surgeries including hemorrhoidectomy, bunionectomy, inguinal hernia repair, total knee arthroplasty, and augmentation mammoplasty. However, like other bupivacaine formulations, the liposomal extended-release bupivacaine does have some side effects. In this brief review, we provide an update of the current knowledge in the use of bupivacaine for postsurgical analgesia.

Mechanical compression of articular cartilage induces chondrocyte proliferation and inhibits proteoglycan synthesis by activation of the ERK pathway: implications for tissue engineering and regenerative medicine.

Articular cartilage is recalcitrant to endogenous repair and regeneration and is thus a focus of tissue engineering and regenerative medicine strategies. A prerequisite for articular cartilage tissue engineering is an understanding of the signal transduction pathways involved in mechanical compression during trauma or disease. We sought to explore the role of the extracellular signal‐regulated kinase 1/2 (ERK 1/2) pathway in chondrocyte proliferation and proteoglycan synthesis following acute mechanical compression. Bovine articular cartilage explants were cultured with and without the ERK 1/2 pathway inhibitor PD98059. Cartilage explants were statically loaded to 40% strain at a strain rate of 1/s for 5 s. Control explants were cultured under similar conditions but were not loaded. There were four experimental groups: (a) no load, without inhibitor; (b) no load, with the inhibitor PD98059; (c) loaded, without the inhibitor; and (d) loaded, with the inhibitor PD98059. The explants were cultured for varying durations from 5 min to 5 days and were then analysed by biochemical and immunohistochemical methods. Mechanical compression induced phosphorylation of ERK 1/2, and this was attenuated with the ERK 1/2 pathway inhibitor PD98059 in a dose‐dependent manner. Chondrocyte proliferation was increased by mechanical compression. This effect was blocked by the inhibitor of the ERK 1/2 pathway. Mechanical compression also led to a decrease in proteoglycan synthesis that was reversed with inhibitor PD98059. In conclusion, the ERK 1/2 pathway is involved in the proliferative and biosynthetic response of chondrocytes following acute static mechanical compression. Copyright

Interleukin-17 receptor-like gene is a novel antiapoptotic gene highly expressed in androgen-independent prostate cancer.

We have recently identified a new gene, interleukin-17 receptor-like (IL-17RL), which is expressed in normal prostate and prostate cancer. This investigation is focused on the role of IL-17RL in prostate cancer. We found that IL-17RL was expressed at significantly higher levels in several androgen-independent prostate cancer cell lines (PC3, DU145, cds1, cds2, and cds3) and tumors compared with the androgen-dependent cell lines (LNCaP and MLC-SV40) and tumors. In an in vivo model of human prostate tumor growth in nude mice (CWR22 xenograft model), IL-17RL expression in tumors was induced by androgen deprivation. The relapsed androgen-independent tumors expressed higher levels of IL-17RL compared with the androgen-dependent tumors. Overexpression of IL-17RL in tumor necrosis factor alpha (TNFalpha)-sensitive LNCaP cells inhibited TNFalpha-induced apoptosis by blocking activation of caspase-3 downstream to caspase-2 and caspase-8. Reciprocally, knocking down IL-17RL expression by small interfering RNA induced apoptosis in all the prostate cancer cell lines studied. Taken together, these results show that IL-17RL is a novel antiapoptotic gene, which may confer partially the property of androgen-independent growth of prostate cancer by promoting cell survival. Thus, IL-17RL is a potential therapeutic target in the treatment of prostate cancer.