Understanding Dendritic Cells and Their Role in Cutaneous Carcinoma and Cancer Immunotherapy

Dendritic cells (DC) represent a diverse group of professional antigen-presenting cells that serve to link the innate and adaptive immune systems. Their capacity to initiate a robust and antigen-specific immune response has made them the ideal candidates for cancer immunotherapies. To date, the clinical impact of DC immunotherapy has been limited, which may, in part, be explained by the complex nature of DC biology. Multiple distinct subsets of DCs have been identified in the skin, where they can be broadly subcategorized into epidermal Langerhans cells (LC), myeloid-derived dermal dendritic cells (mDC) and plasmacytoid dendritic cells (pDC). Each subset is functionally unique and may activate alternate branches of the immune system. This may be relevant for the treatment of squamous cell carcinoma, where we have shown that the tumor microenvironment may preferentially suppress the activity of mDCs, while LCs remain potent stimulators of immunity. Here, we provide an in depth analysis of DC biology, with a particular focus on skin DCs and their role in cutaneous carcinoma. We further explore the current approaches to DC immunotherapy and provide evidence for the targeting of LCs as a promising new strategy in the treatment of skin cancer

Transforming growth factor-β receptor types I and II are expressed in renal tubules and are increased after chronic unilateral ureteral obstruction

Transforming growth factor-β (TGF-β) is a profibrotic cytokine which has been implicated in the renal fibrosis which follows unilateral ureteral obstruction (UUO) in the rat. TGF-β receptor types I (TGF-RI) and TGF-β. We sought to determine if TGF-Ri and TGF-RII are found in the kidney, and if their expression is changed as a result of UUO. Polymerase chain reaction (PCR) was used to determine expression of mRNA for TGF-RI and TGF-RII in the kidney. Immunoperoxidase was used to localize and quantify the expression of these receptors at 3, 7, 14, 21 and 28 days after UUO, and in sham-operated animals. Expression of mRNA for TGF-RI and TGF-RII was demonstrated in sham operated, obstructed ad contralateral unobstructed kidneys using PCR. Using immunoperoxidase, a uniform kidneys, whereas medullary tubules showed a patchy TGF-RI distribution and no TGF-RII staining. After UUO, an increased tubular expression of TGF-RI and TGF-RII was noted in both obstructed and contralateral kidneys compared to sham operated kidneys. No staining for either TGF-RI or TGF- RII was noted in glomeruli, vasculature or intestinal cells. TGF-β receptors I and II were found exclusively in renal tubules and were shown to increase in both the obstructed and contralateral kidneys relative to sham operated animals. Upregulation of TGF-β receptors in both kidneys suggest that TGF-β may contribute to the fibrotic response in the obstructed kidney and the hypertrophic response of the contralateral kidney

Chronic unilateral ureteral obstruction is associated with interstitial fibrosis and tubular expression of transforming growth factor-β

Chronic unilateral ureteral obstruction (UUO) results in interstitial fibrosis and nephron damage associated with irreversible loss of function. Collagen is increased in UUO, but detailed studies of rat renal extracellular matrix changes in UUO have not been carried out. Acute (3-day) obstruction results in increases in renal macrophages and the fibrogenic cytokine transforming growth factor-β (TGF-β), but their involvement in longer-term obstruction and fibrosis has not been studied. In the present experiments, kidneys of rats after UUO of 0, 1, 2, 3, 7, 14, 21, and 28 days\u27 duration were used. Trichrome staining, measurement of interstitial volume, and immunohistochemical studies localizing collagens I, III, and IV; laminin; fibronectin; TGF-β; and macrophages were carried out. We found increases in the interstitial space in both codex and medulla that (a) were significant by day 7 after UUO and (b) were accompanied by increased deposition of collagen I and collagen III. Collagen IV, laminin, and fibronectin, normally associated with the basement membrane, were found both in a thickened basement membrane and in the interstitial space. Macrophages, not found in sham-operated kidneys, were found in the interstitial space after UUO. TGF- β was found in sham cortical tubules, but not in medullary tubules. UUO was associated with little change in cortical TGF-β, whereas at 14 days, TGF-β was found in dilated medullary tubules. Immunohistochemical findings were confirmed with measurements of tissue TGF-β. In summary, UUO is associated with interstitial fibrosis. The increase in extracellular matrix is due both to increases in the interstitial collagens I and III and the basement membrane-associated collagen IV, laminin, and fibronectin. Macrophages are increased after UUO, but do not seem to be associated with the fibrogenic cytokine TGF-β. Medullary tubular synthesis of TGF-β may be a contributing factor in the fibrosis associated with UUO

MAGE-A3 is a prognostic biomarker for poor clinical outcome in cutaneous squamous cell carcinoma with perineural invasion via modulation of cell proliferation.

Perineural invasion is a pathologic process of neoplastic dissemination along and invading into the nerves. Perineural invasion is associated with aggressive disease and a greater likelihood of poor outcomes. In this study, 3 of 9 patients with cutaneous squamous cell carcinoma and perineural invasion exhibited poor clinical outcomes. Tumors from these patients expressed high levels of MAGE-A3, a cancer testis antigen that may contribute to key processes of tumor development. In addition to perineural invasion, the tumors exhibited poor differentiation and deep invasion and were subsequently classified as Brigham and Women's Hospital tumor stage 3. Cyclin E, A and B mRNA levels were increased in these tumors compared with normal skin tissues (102.93±15.03 vs. 27.15±4.59, 36.83±19.41 vs. 11.59±5.83, 343.77±86.49 vs. 95.65±29.25, respectively; p<0.05). A431 cutaneous squamous cell carcinoma cells pretreated with MAGE-A3 antibody exhibited a decreased percentage S-phase cells (14.13±2.8% vs. 33.97±1.1%; p<0.05) and reduced closure in scratch assays (43.88±5.49% vs. 61.17±3.97%; p = 0.0058). In a syngeneic animal model of squamous cell carcinoma, immunoblots revealed overexpression of MAGE-A3 and cyclin E, A, and B protein in tumors at 6 weeks. However, knockout of MAGE-A3 expression caused a reduction in tumor growth (mean tumor volume 155.3 mm3 vs. 3.2 mm3) compared with parental cells. These results suggest that MAGE-A3 is a key mediator in cancer progression. Moreover, elevated collagen XI and matrix metalloproteases 3, 10, 11, and 13 mRNA levels were observed in poorly differentiated cutaneous squamous cell carcinoma with perineural invasion compared with normal skin tissue (1132.56±882.7 vs. 107.62±183.62, 1118.15±1109.49 vs. 9.5±5, 2603.87±2385.26 vs. 5.29±3, 957.95±627.14 vs. 400.42±967.66, 1149.13±832.18 vs. 19.41±35.62, respectively; p<0.05). In summary, this study highlights the potential prognostic value of MAGE-A3 in clinical outcomes of cutaneous squamous cell carcinoma patients

Multiphoton Imaging and Laser Ablation of Rodent Spermatic Cord Nerves: Potential Treatment for Patients With Chronic Orchialgia

Microsurgical denervation of the spermatic cord has been done to treat chronic orchialgia. However, identifying the site of spermatic cord nerves is not feasible with an operating microscope or robotic stereoscope. We used multiphoton microscopy, a novel laser imaging technology, to identify and selectively ablate spermatic cord nerves in the rat. The spermatic cords of adult male Sprague-Dawley® rats were initially imaged in vivo under a low power multiphoton microscopy laser. After assessing the number, diameter and site (vasal vs perivasal) of the nerves a higher power laser using the same objective was used to ablate the nerves. The precision of nerve ablation and the preservation of surrounding structures were determined by histological analysis. We assessed the heterogeneity of the number of nerves with the Wilcoxon signed rank test. The average number of nerves per spermatic cord was 10, which was similar bilaterally (p = 0.13). The vas and perivasal structures had a similar number of nerves (p = 0.4). The median diameter of all nerves was 32 μm. Confirmation of nerve ablation, and preservation of the vas deferens and vasculature were anatomically validated by histological analysis. Multiphoton microscopy can identify and ablate nerves selectively in vivo in the rat. It can potentially be used for spermatic cord denervation to treat chronic orchialgia. Such imaging may increase the efficacy of nerve ablation and can avoid the potential risks of testicular atrophy and hydrocele associated with spermatic cord microsurgical denervation