Yoshiko Watanabe

Tumor-infiltrating lymphocytes, particularly the balance between CD8+ T cells and CCR4+ regulatory T cells, affect the survival of patients with oral squamous cell carcinoma

OBJECTIVE The objective of this study was to clarify the prognostic significance of tumor-infiltrating lymphocytes (TILs) in oral squamous cell carcinoma (OSCC); the present study analyzed various TIL-related parameters. STUDY DESIGN Immunohistochemistry was performed in 87 patients with OSCC for the following TIL-related parameters: nest-CD8(+) T cells, stromal CD8(+) T cells, CD4(+) T cells, total regulatory T cells (Tregs), CCR4(+) Tregs, ratio of nest CD8(+) T cells/CCR4(+) Tregs, and ratio of stromal CD8(+) T cells/CCR4(+) Tregs. RESULTS In univariate analyses, the following parameters were associated with decreased survival: few nest- and stromal CD8(+) T cells and more stromal CCR4(+) Tregs, but not total Tregs. Low ratios of nest and stromal CD8(+) T cell/CCR4(+) Treg were associated with worse survival. In multivariate analysis, the stromal CD8(+) T cell/CCR4(+) Treg ratio was an independent prognostic factor. CONCLUSION Host immune responses in the stroma of OSCC affect the survival of the patients. The in situ balance between effector T cells and regulatory T cells is the most important factor predicting survival.

Eotaxin-3/CC Chemokine Ligand 26 Is a Functional Ligand for CX3CR1

Eotaxin-3/CCL26 is a functional ligand for CCR3 and abundantly produced by IL-4–/IL-13–stimulated vascular endothelial cells. CCL26 also functions as a natural antagonist for CCR1, CCR2, and CCR5. In this study, we report that CCL26 is yet a functional ligand for CX3CR1, the receptor for fractalkine/CX3CL1, which is expressed by CD16+ NK cells, cytotoxic effector CD8+ T cells, and CD14lowCD16high monocytes. Albeit at relatively high concentrations, CCL26 induced calcium flux and chemotaxis in mouse L1.2 cells expressing human CX3CR1 but not mouse CX3CR1 and competed with CX3CL1 for binding to CX3CR1. In chemotaxis assays using human PBMCs, CCL26 attracted not only eosinophils but also CD16+ NK cells, CD45RA+CD27−CD8+ T cells, and CD14lowCD16high monocytes. Intraperitoneal injection of CCL26 into mice rapidly recruited mouse eosinophils and intravenously transferred human CD16+ NK cells into the peritoneal cavity. IL-4–stimulated HUVECs produced CCL26 and efficiently induced adhesion of cells expressing CX3CR1. Real-time PCR showed that skin lesions of psoriasis consistently contained CX3CL1 mRNA but not CCL26 mRNA, whereas those of atopic dermatitis contained CCL26 mRNA in all samples but CX3CL1 mRNA in only about half of the samples. Nevertheless, the skin lesions from both diseases consistently contained CX3CR1 mRNA at high levels. Thus, CCL26 may be partly responsible for the recruitment of cells expressing CX3CR1 in atopic dermatitis particularly when the expression of CX3CL1 is low. Collectively, CCL26 is another agonist for CX3CR1 and may play a dual role in allergic diseases by attracting eosinophils via CCR3 and killer lymphocytes and resident monocytes via CX3CR1.

Differing Phenotypes between Intraepithelial and Stromal Lymphocytes in Early-Stage Tongue Cancer

The significance of tumor-infiltrating lymphocytes (TIL) has attracted much attention in relation to the prognosis of patients. We herein examined the activation status of the TILs in relation to the tumor microenvironment. By using frozen sections of human early-stage tongue cancers (n = 22), the TILs in the cancer nests and those in the cancer stroma were compared for the expression of PD-1, NKG2A, NKG2D, CD69, and Ki-67. The lymphocytes in oral lichen planus, an active immune response-mediated mucosal disease, were also analyzed for comparison purposes. All of the cancer specimens were abundantly infiltrated by CD8(+) T cells and CD56(+) natural killer (NK) cells in the stroma, as well as in the tumor nest. The tumor nest-infiltrating (intraepithelial) CD8(+) T cells frequently expressed PD-1, an inhibitory receptor, in sharp contrast to those in the stroma or in the lichen planus. Conversely, the intraepithelial CD8(+) T cells only infrequently expressed NKG2D, an activating receptor, in contrast to those in the stroma or in the lichen planus. No intraepithelial CD8(+) T cells expressed Ki-67, a proliferation-associated marker, whereas those in the stroma frequently expressed it. Furthermore, the intraepithelial NK cells expressed NKG2A, an inhibitory receptor, more frequently than those in the stroma or the lichen planus. Collectively, the intraepithelial CD8(+) T cells and NK cells are phenotypically inactivated, whereas stromal counterparts are phenotypically just as active as those in the lichen planus. These results suggest the first-step occurrence of an immune evasion mechanism in the tumor nest of oral squamous cell carcinoma.

Imaging mass spectrometry distinguished the cancer and stromal regions of oral squamous cell carcinoma by visualizing phosphatidylcholine (16:0/16:1) and phosphatidylcholine (18:1/20:4)

Most oral cancers are oral squamous cell carcinoma (OSCC). The anatomical features of OSCC have been histochemically evaluated with hematoxylin and eosin. However, the border between the cancer and stromal regions is unclear and large portions of the cancer and stromal regions are resected in surgery. To reduce the resected area and maintain oral function, a new method of diagnosis is needed. In this study, we tried to clearly distinguish the border on the basis of biomolecule distributions visualized by imaging mass spectrometry (IMS). In the IMS dataset, eleven signals were significantly different in intensity (p < 0.01) between the cancer and stromal regions. Two signals at m/z 770.5 and m/z 846.6 were distributed in each region, and a clear border was revealed. Tandem mass spectrometric (MS/MS) analysis identified these signals as phosphatidylcholine (PC) (16:0/16:1) at m/z 770.5 in the cancer region and PC (18:1/20:4) at m/z 846.6 in the stromal region. Moreover, the distribution of PC species containing arachidonic acid in the stromal region suggests that lymphocytes accumulated in response to the inflammation caused by cancer invasion. In conclusion, the cancer and stromal regions of OSCCs were clearly distinguished by use of these PC species and IMS analysis, and this molecular identification can provide important information to elucidate the mechanism of cancer invasion.

Improved technique for evaluating oral free flaps by pinprick testing assisted by indocyanine green near-infrared fluorescence angiography

In head and neck surgery, free-flap reconstruction using a microvascular anastomosis is an indispensable option after tumor ablation. Because the success of free-flap reconstruction is enhanced by rapid identification and salvage of failing flaps, postoperative monitoring of free flaps is essential. We describe a new technique using indocyanine green (ICG) near-infrared angiography and pinprick testing to monitor intraoral free flaps. A solution of ICG (Diagnogreen, 5 ml) was intravenously injected, and scanning was performed with a near-infrared video camera system. Thirty seconds after ICG injection, a pinprick test was performed by placing a 24-gage needle through the dermis to the subcutaneous fat of the flap. Pinprick testing during ICG fluorescence imaging was performed in 30 patients. Flap perfusion was confirmed in all patients, and all flaps survived postoperatively. ICG fluorescence imaging demonstrated that flap perfusion was maintained.

Matrix-assisted laser desorption/ionization imaging mass spectrometry revealed traces of dental problem associated with dental structure

Periodontal disease is a serious dental problem because it does not heal naturally and leads to tooth loss. In periodontal disease, inflammation at periodontal tissue is thought as predominant, and its effect against tooth itself remains unclear. In this study, we applied matrix-assisted laser desorption/ionization imaging mass spectrometry (IMS) to teeth for the first time. By comparing anatomical structure of tooth affected with periodontal disease with normal ones, we analyzed traces of the disease on tooth. We found signals characteristic of enamel, dentin, and dental pulp, respectively, in mass spectra obtained from normal teeth. Ion images reconstructed using these signals showed anatomical structures of the tooth clearly. Next, we performed IMS upon teeth of periodontal disease. Overall characteristic of the mass spectrum appeared similar to normal ones. However, ion images reconstructed using signals from the tooth of periodontal disease revealed loss of periodontal ligament visualized together with dental pulp in normal teeth. Moreover, ion image clearly depicted an accumulation of signal at m/z 496.3 at root surface. Such an accumulation that cannot be examined only from mass spectrum was revealed by utilization of IMS. Recent studies about inflammation revealed that the signal at m/z 496.3 reflects lyso-phosphatidylcholine (LPC). Infiltration of the signal is statistically significant, and its intensity profile exhibited the influence has reached deeply into the tooth. This suggests that influence of periodontal disease is not only inflammation of periodontal tissue but also infiltration of LPC to root surface, and therefore, anti-inflammatory treatment is required besides conventional treatments.

End-to-side anastomosis to the external jugular vein: preservation of external jugular vein blood flow

v p the anastomosis of the artery, the vein of the donor flap is anastomosed to the external jugular vein. The anastomosis is he external jugular vein provides a long, free vessel that acilitates microsurgical anastomosis in free tissue transfers f the head and neck. However, it is liable to kink because of ts length, which increases the risk of venous thrombosis.1,2 arly in a conventional neck dissection the vein is identied and ligated near the inferior pole of the parotid gland. hen a microvascular free flap is to be used for recontruction, the entire external jugular vein is dissected and reserved with a suture-ligation at the upper end. Because f the prolonged duration of ischaemia before revascularisaion, clots can form in its lumen and may be found at the time f anastomosis. Even after revascularisation using an end-tond anastomosis, the decreased venous flow may accelerate enous thrombosis.3 We describe an end-to-side anastomois to the external jugular vein that preserves blood flow by voiding ligation and decreases the likelihood of thrombosis.

Three-dimensional computed tomographic analysis of variations of the carotid artery.

BACKGROUND Although the terms tortuous, coiling, and kinking have been used to describe the curvature of the carotid artery, the prevalence rates of these patterns have differed among studies. We morphologically evaluated the characteristics of the carotid artery by means of three-dimensional computed tomography (3DCT) to clarify the prevalence of tortuosity, coiling, and kinking. We present our results and discuss the clinical impact of our findings. METHODS A total 148 patients underwent contrast-enhanced CT (including 55 patients who underwent dynamic CT), and anatomical variations were analyzed on the basis of 3DCT images. RESULTS Among the 296 arteries, tortuosity was present in 254 (85.8%), coiling in 9 (3.0%), kinking in 3 (1.0%), and occlusion in 2 (0.7%). CONCLUSION 3DCT image reconstruction is an effective means for classifying morphological variations of the ICA and detecting abnormalities of the carotid artery. It can thereby potentially reduce the risk of serious complications during neck surgery.

Synchronous Repair of Bilateral Cleft Lip in a Patient with Wolf-Hirschhorn (4p-) Syndrome

Abstract This report is of an infant with Wolf-Hirschhorn (4p-) syndrome, a disorder of multiple malformations that is characterised by delayed growth and development, and facial dysmorphism. The bilateral cleft lip was successfully repaired by synchronous repair surgery at age 1 year 5 months.