Platelet-dependent arterial thrombosis triggers most heart attacks and strokes. Because the coagulation protease thrombin is the most potent activator of platelets, identification of the platelet receptors for thrombin is critical for understanding thrombosis and haemostasis. Protease-activated receptor-1 (PAR1) is important for activation of human platelets by thrombin, but plays no apparent role in mouse platelet activation. PAR3 is a thrombin receptor that is expressed in mouse megakaryocytes. Here we report that thrombin responses in platelets from PAR3-deficient mice were markedly delayed and diminished but not absent. We have also identified PAR4, a new thrombin-activated receptor. PAR4 messenger RNA was detected in mouse megakaryocytes and a PAR4-activating peptide caused secretion and aggregation of PAR3-deficient mouse platelets. Thus PAR3 is necessary for normal thrombin responses in mouse platelets, but a second PAR4-mediated mechanism for thrombin signalling exists. Studies with PAR-activating peptides suggest that PAR4 also functions in human platelets, which implies that an analogous dual-receptor system also operates in humans. The identification of a two-receptor system for platelet activation by thrombin has important implications for the development of antithrombotic therapies.

Secondary lymphoid organ chemokine (SLC) is expressed in high endothelial venules and in T cell zones of spleen and lymph nodes (LNs) and strongly attracts naive T cells. In mice homozygous for the paucity of lymph node T cell (plt) mutation, naive T cells fail to home to LNs or the lymphoid regions of spleen. Here we demonstrate that expression of SLC is undetectable in plt mice. In addition to the defect in T cell homing, we demonstrate that dendritic cells (DCs) fail to accumulate in spleen and LN T cell zones of plt mice. DC migration to LNs after contact sensitization is also substantially reduced. The physiologic significance of these abnormalities in plt mice is indicated by a markedly increased sensitivity to infection with murine hepatitis virus. The plt mutation maps to the SLC locus; however, the sequence of SLC introns and exons in plt mice is normal. These findings suggest that the abnormalities in plt mice are due to a genetic defect in the expression of SLC and that SLC mediates the entry of naive T cells and antigen-stimulated DCs into the T cell zones of secondary lymphoid organs.

Preferential homing of naive lymphocytes to secondary lymphoid organs is thought to involve the action of chemokines, yet no chemokine has been shown to have either the expression pattern or the activities required to mediate this process. Here we show that a chemokine represented in the EST database, secondary lymphoid-tissue chemokine (SLC), is expressed in the high endothelial venules of lymph nodes and Peyer's patches, in the T cell areas of spleen, lymph nodes, and Peyer's patches, and in the lymphatic endothelium of multiple organs. SLC is a highly efficacious chemoattractant for lymphocytes with preferential activity toward naive T cells. Moreover, SLC induces firm adhesion of naive T lymphocytes via ␤2 integrin binding to the counter receptor, intercellular adhesion molecule-1, a necessary step for lymphocyte recruitment. SLC is the first chemokine demonstrated to have the characteristics required to mediate homing of lymphocytes to secondary lymphoid organs. In addition, the expression of SLC in lymphatic endothelium suggests that the migration of lymphocytes from tissues into efferent lymphatics may be an active process mediated by this molecule.

Activated T lymphocytes differentiate into effector cells tailored to meet disparate challenges to host integrity. For example, type 1 and type 2 helper (T(H)1 and T(H)2) cells secrete cytokines that enhance cell-mediated and humoral immunity, respectively. The chemokine monocyte chemoattractant protein-1 (MCP-1) can stimulate interleukin-4 production and its overexpression is associated with defects in cell-mediated immunity, indicating that it might be involved in T(H)2 polarization. Here we show that MCP-1-deficient mice are unable to mount T(H)2 responses. Lymph node cells from immunized MCP-1(-/-) mice synthesize extremely low levels of interleukin-4, interleukin-5 and interleukin-10, but normal amounts of interferon-gamma and interleukin-2. Consequently, these mice do not accomplish the immunoglobulin subclass switch that is characteristic of T(H)2 responses and are resistant to Leishmania major. These effects are direct rather than due to abnormal cell migration, because the trafficking of naive T cells is undisturbed in MCP-1(-/-) mice despite the presence of MCP-1-expressing cells in secondary lymphoid organs of wild-type mice. Thus, MCP-1 influences both innate immunity, through effects on monocytes, and adaptive immunity, through control of T helper cell polarization.

Her-2-neu expression appears to increase with progression to androgen independence. Thus, therapeutic targeting of this tyrosine kinase in prostate cancer may be warranted.

Periostin is a recently identified gene that is preferentially expressed in periosteum, indicating a potential role in bone formation and maintenance of structure. We independently identified and isolated periostin from cancer tissue, using the palindromic PCR-driven cDNA Differential Display technique. For the present work, we developed a novel sandwich chemiluminescence assay to detect serum periostin level using newly developed monoclonal and polyclonal antibodies. We investigated serum periostin levels in breast cancer and small cell lung cancer patients, especially in patients with bone metastasis. The study included 58 breast cancer and 44 small cell lung cancer patients. Serum periostin levels were elevated in breast cancer patients presenting with bone metastases (92.0 +/- 28.6 ng/ml) compared to similar breast cancer patients without evidence of bone metastasis (55.0 +/- 16.6 ng/ml, p = 0.04). No correlation was found between the serum periostin level and any other prognostic factors, such as clinical stage and lymph node metastasis in breast cancer. Serum periostin levels thus appear to serve as a marker of bone metastasis from breast cancer. In contrast, serum periostin levels were similar in samples from patients with small cell lung cancer who did or did not have bone metastasis. However, increasing T-stage and N-stage of patients with small cell lung cancer were correlated with higher periostin levels (T4, 126.5 +/- 29.7 ng/ml v.s. T2, 64.9 +/- 16.1 ng/ml, p = 0.03; and T4 v.s. T1, 36.3+/- 7.5 ng/ml, p = 0.01; N3, 108.7 +/- 17.3 ng/ml v.s. N2, 49.7+/- 10.9 ng/ml, p = 0.01). Periostin has a substantial homology with the insect cell adhesion molecule, fasciclin I. Thus, expression of periostin may facilitate tumor cell adhesion to the bone surface. In fact, we found by in situ RNA hybridization, that the periostin gene was highly expressed in the stromal cells immediately surrounding the tumor, but not within the breast cancer cells themselves.

These studies strongly suggest that the heart synthesizes and secretes apoB-containing lipoproteins.

We used our palindromic polymerase chain reaction (PCR)-driven cDNA differential display technique to identify and isolate a gene, designated periostin, from cancer tissues and found it to be overexpressed in several human tumors. We attempted to determine the influence of periostin expression on clinical outcome in patients with non-small cell lung cancer (NSCLC) by reverse transcriptase (RT)-PCR analysis. Periostin gene was highly expressed at the tumor periphery of lung cancer tissue but not within the tumor by in situ RNA hybridization, suggesting that expression of periostin may be involved in the process of tumor invasion. Periostin transcripts were detected in 50 (49.0%) of the tumor samples, although some paired normal lung samples showed weak expression. There was no relationship between periostin gene expression and gender, N-or Tstatus. The NSCLC patients with periostin expression had significantly poorer survival than the patients without periostin expression (P = = = =0.0338). Key words: Periostin-RT-PCR-Lung cancer-PrognosisLung cancer is a major cause of death from malignant diseases, due to its high incidence and malignant behavior, and the lack of major advances in treatment strategy. 1)Lung cancer was the leading indication for respiratory surgery (42.2%) in 1998 in Japan, with more than 15 000 patients undergoing surgical operation at Japanese institutions.2) The clinical behavior of non-small lung cancer (NSCLC) is largely associated with its stage.3) However, a significant number of tumors in patients with early-stage NSCLC show aggressive behavior and a tendency to relapse. Moreover, the prognosis for patients with operable NSCLC remains gloomy in comparison with that observed in operable gastric, colon, or breast cancer.4) A variety of clinicopathologic characteristics may affect prognosis. 5)Despite steadily accumulating evidence that numerous genetic markers influence the biological behavior of NSCLC, the intrinsic nature of the gene deregulation that leads small tumors to metastasize remains highly elusive. 6)The abnormal expression of certain genes in cancer cells is closely related to various aspects of tumor progression, including tumor growth, invasion and metastasis. Recently, we used the palindromic polymerase chain reaction (PCR)-driven cDNA differential display technique developed in our laboratory 7) to identify and isolate periostin cDNA, and found it to be overexpressed in several human tumors including primary colon cancer, metastatic liver cancer from colon cancer and breast cancer (Bao et al., manuscript in preparation). The periostin protein shares structural and sequence homology with fasciclin I, which is an insect adhesion molecule.8-10) It was reported that cellular adhesion molecules play an important role in the process of metastasis. 11)As available prognostic markers leave much to be desired for NSCLC, we investigated periostin transcript in patients with NSCLC by means of a reverse transcriptase PCR (RT-PCR) and in situ hybridization analysis, and we assessed t...