The firm adhesion and transplatelet migration of leukocytes on vascular thrombus are both dependent on the interaction of the leukocyte integrin, Mac-1, and a heretofore unknown platelet counterreceptor. Here, we identify the platelet counterreceptor as glycoprotein (GP) Ibα, a component of the GP Ib-IX-V complex, the platelet von Willebrand factor (vWf) receptor. THP-1 monocytic cells and transfected cells that express Mac-1 adhered to GP Ibα–coated wells. Inhibition studies with monoclonal antibodies or receptor ligands showed that the interaction involves the Mac-1 I domain (homologous to the vWf A1 domain), and the GP Ibα leucine-rich repeat and COOH-terminal flanking regions. The specificity of the interaction was confirmed by the finding that neutrophils from wild-type mice, but not from Mac-1–deficient mice, bound to purified GP Ibα and to adherent platelets, the latter adhesion being inhibited by pretreatment of the platelets with mocarhagin, a protease that specifically cleaves GP Ibα. Finally, immobilized GP Ibα supported the rolling and firm adhesion of THP-1 cells under conditions of flow. These observations provide a molecular target for disrupting leukocyte–platelet complexes that promote vascular inflammation in thrombosis, atherosclerosis, and angioplasty-related restenosis.

No abstract

Orthotopic liver transplantation (OLT) is the only curative therapy of HCC with underlying cirrhosis, but due to HCC metastasis and recurrence, its benefit is limited to a small population who meet the strict selection criteria. We previously reported that Licartin ([ 131 I]mAb HAb18G/CD147) was safe and effective in treating HCC patients, and its antigen, HAb18G/ CD147, was closely related to HCC invasion and metastasis. Here, we reported a randomized controlled trial to assess the post-OLT antirecurrence efficacy of Licartin in advanced HCC patients. We randomized 60 post-OLT patients with HCC, who were at tumor stage 3/4 and outside the Milan criteria before OLT, into 2 groups. Three weeks after OLT, the treatment group received 15.4 MBq/kg of Licartin, while the control group received placebo intravenously for 3 times with an interval of 28 days. At 1-year follow-up, the recurrence rate significantly decreased by 30.4% (P ‫؍‬ 0.0174) and the survival rate increased by 20.6% (P ‫؍‬ 0.0289) in the treatment group, compared with those in the control group. For the control group versus the treatment group, the hazard ratio for recurrence H epatocellular carcinoma is the most common type of primary liver cancer and ranks sixth among cancers as a cause of death worldwide. 1 It is a highly malignant tumor characterized by rapid progression, poor prognosis, and frequent tumor recurrence. It has an annual incidence rate of 564,000 cases, and 55% of those are in China. 2 The mean natural survival time was reported to be only 3-6 months due to the rapid progression of tumor, especially the spread and metastasis. 3,4 Surgery is the preferred treatment, but less than 20% of patients have the chance to be treated surgically Abbreviations: AFP, alpha fetoprotein; CI, confidence interval; DBIL, direct bilirubin; mAb, monoclonal antibody; OLT, orthotopic liver transplantation; TNM, tumor-nodes-metastasis. From the

CD147 molecule is reported to be correlated with the malignancy of some cancers; however, it remains unclear whether it is involved in the progression of hepatocellular carcinoma (HCC). Here, we investigated the function of HAb18G/CD147, a member of CD147 family, and its antibodies, HAb18 and LICARTIN, in HCC invasion and metastasis. We observed that HAb18G/CD147 gene silence in HCC cells significantly decreased the secretion of matrix metalloproteinase (MMP) and the invasive potential of HCC cells (P < 0.001). MMP silence in HCC cells also significantly suppressed the invasion of the cells when cocultured with fibroblasts; however, its inhibitory effect was significantly weaker than that of both HAb18G/CD147 silence in HCC cells and that of MMP silence in fibroblasts (P < 0.001). Blocking the HAb18G/CD147 molecule on HCC cells with HAb18 monoclonal antibody resulted in a similar suppressive effect on MMP secretion and cell invasion, but with no significant effects on the cell growth. 131 I-labeled HAb18 F(ab ¶) 2 (LICARTIN), however, significantly inhibited the in vitro growth of HCC cells (P < 0.001). In an orthotopic model of HCC in nude mice, HAb18 and LICARTIN treatment effectively reduced the tumor growth and metastasis as well as the expression of three major factors in the HCC microenviroment (MMPs, vascular endothelial growth factor, and fibroblast surface protein) in the paracancer tissues. Overall, these results suggest that HAb18G/CD147 plays an important role in HCC invasion and metastasis mainly via modulating fibroblasts, as well as HCC cells themselves to disrupt the HCC microenviroment. LICARTIN can be used as a drug targeting to HAb18G/ CD147 in antimetastasis and recurrence therapy of HCC.

Connexin (Cx) 43 serves important roles in bone function and development. Targeted deletion of Cx43 in osteoblasts or osteocytes leads to increased osteocyte apoptosis, osteoclast recruitment, and reduced biomechanical properties. Cx43 forms both gap junction channels and hemichannels, which mediate the communication between adjacent cells or between cell and extracellular environments, respectively. Two transgenic mouse models driven by a DMP1 promoter with the overexpression of dominant negative Cx43 mutants were generated to dissect the functional contribution of Cx43 gap junction channels and hemichannels in osteocytes. The R76W mutant blocks gap junction channel, but not hemichannel function, and the Δ130-136 mutant inhibits activity of both types of channels. Δ130-136 mice showed a significant increase in bone mineral density compared to WT and R76W mice. MicroCT analyses revealed a significant increase in total tissue and bone area in midshaft cortical bone of Δ130-136 mice. The bone marrow cavity was expanded, whereas the cortical thickness was increased and associated with increased bone formation along the periosteal area. However, there is no significant alteration in the structure of trabecular bone. Histologic sections of the midshaft showed increased apoptotic osteocytes in Δ130-136, but not in WT and R76W, mice which correlated with altered biomechanical and estimated bone material properties. Osteoclasts were increased along the endocortical surface in both transgenic mice with a greater effect in Δ130-136 mice which likely contributed to the increased marrow cavity. Interestingly, the overall expression of serum bone formation and resorption markers were higher in R76W mice. These findings suggest that osteocytic Cx43 channels play distinctive roles in the bone; hemichannels play a dominant role in regulating osteocyte survival, endocortical bone resorption and periosteal apposition, and gap junction communication is involved in the process of bone remodeling.

Recent clinical trials suggest that blockade of integrins is a promising strategy for the treatment of acute coronary syndromes. Administration of 7E3 monoclonal antibody (mAb) Fab fragment (c7E3 Fab) directed against platelet integrin IIb/IIIa (alpha IIb beta 3, CD41/CD61) reduces acute ischemic complications of coronary angioplasty and clinical restenosis at 6 months. However, 7E3 mAb is not selective for platelet IIb/IIIa but also cross-reacts with the leukocyte integrin Mac-1 (alpha M beta 2, CD11b/CD18) and the vitronectin receptor (alpha v beta 3, CD51/CD61). Information regarding how this mAb may affect other cells important in vascular repair is scant. Potential interactions of c7E3 Fab with inflammatory (i.e., monocytes and neutrophils), vascular smooth muscle, and endothelial cells may contribute to the in vivo actions of c7E3 Fab. In this study we explored the binding of 7E3 to monocytic cells and the functional effect of 7E3 and c7E3 Fab on Mac-1-mediated adhesion to fibrinogen (FGN) and intercellular adhesion molecule-1 (ICAM-1), ligands abundant in the injured vessel wall. Flow cytometry demonstrated that 7E3 bound to THP-1 monocytic cells and identified a subpopulation (approximately 10%) of Mac-1 that was qualitatively similar to that recognized by CBRM1/5, a mAb directed to an activation-specific neoepitope present on a subset of Mac-1 molecules. mAb 7E3 bound to K562 cells transfected with just the alpha subunit (CD11b) of Mac-1 but not to nontransfected cells, confirming a direct interaction between 7E3 and Mac-1. mAb 7E3 and c7E3 Fab blocked the adhesion of Mac-1-bearing cells to FGN (80 +/- 11% and 78 +/- 9% inhibition, respectively) and ICAM-1 (62 +/- 14% and 62 +/- 17%). Both 7E3 and c7E3 Fab significantly inhibited (70 +/- 6% and 62 +/- 26%) soluble FGN binding to human peripheral blood monocytes. Thus, c7E3 Fab cross-reacts with the CD11b subunit of Mac-1 and interrupts cell-extracellular matrix and cell-cell adhesive interactions and may thereby influence the recruitment of circulating monocytes to sites of vessel injury. Given the recent evidence that adherent and infiltrating monocyte number directly correlates with the extent of neointimal hyperplasia, inhibition of Mac-1-dependent adhesion and IIb/IIIa-dependent function by c7E3 Fab may jointly contribute to the regulation of vascular repair and to the sustained clinical benefits observed with c7E3 Fab after angioplasty.

The leukocyte integrin Mac-1 (CD11b/CD18) and the urokinase receptor (uPAR, CD87) mediate complementary functions in myelomonocytic cells. Both receptors promote degradation of fibrin(ogen) and also confer adhesive properties on cells because Mac-1 and uPAR bind fibrin and vitronectin, respectively. Staining of lung biopsy specimens from patients with acute lung injury indicated that fibrin and vitronectin colocalize at exudative sites in which macrophages bearing these receptors accumulate. Because of the parallel roles and physical proximity of Mac-1 and uPAR, the capacity of these receptors to functionally interact was explored. Induction of Mac-1 and uPAR expression on monocytic cell lines by transforming growth factor-beta 1 and 1.25-(OH)2 vitamin D3 conferred urokinase and uPAR-dependent adhesion to vitronectin, which was further promoted by engagement of Mac-1. Vitronectin attachment promoted subsequent Mac-1-mediated fibrinogen degradation threefold to fourfold. In contrast, enhancement of uPAR occupancy by exogenous urokinase or receptor binding fragments thereof inhibited Mac-1 function. Addition of urokinase progressively inhibited Mac-1-mediated fibrinogen binding and degradation (maximal inhibition, 91% +/- 14% and 72% +/- 15%, respectively). Saturation of uPAR with urokinase also inhibited binding of the procoagulant Mac-1 ligand, Factor X. These inhibitory effects of uPAR were reproduced in fresh monocytes, cultured monocytic cells, and in Chinese hamster ovary (CHO) cells transfected with both human Mac-1 and human uPAR. These data show that the procoagulant and fibrinolytic potential of monocytic cells is co-ordinately regulated by ligand binding to both Mac-1 and uPAR and identify uPAR as a regulator of integrin function. Vitronectin-enhanced fibrin(ogen) turnover by Mac-1 may operate as a salvage pathway in the setting of urokinase and plasmin inhibitors to promote clearance of the provisional matrix and subsequent healing.

No abstract