The CCN family comprises cysteine-rich 61 (CYR61/CCN1), connective tIssue growth factor (CTGF/CCN2), nephroblastoma overexpressed (NOV/CCN3), and Wnt-induced secreted proteins-1 (WISP-1/CCN4), -2 (WISP-2/CCN5) and -3 (WISP-3/CCN6). These proteins stimulate mitosis, adhesion, apoptosis, extracellular matrix production, growth arrest and migration of multiple cell types. Many of these activities probably occur through the ability of CCN proteins to bind and activate cell surface integrins. Accumulating evidence supports a role for these factors in endocrine pathways and endocrine-related processes. To illustrate the broad role played by the CCN family in basic and clinical endocrinology, this Article highlights the relationship between CCN proteins and hormone action, skeletal growth, placental angiogenesis, IGF-binding proteins and diabetes-induced fibrosis.

CYR61 (CCN1) is a member of the CCN family of secreted matricellular proteins that includes connective tissue growth factor (CCN2), NOV (CCN3), WISP-1 (CCN4), WISP-2 (CCN5), and WISP-3 (CCN6). First identified as the product of a growth factor-inducible immediate-early gene, CYR61 is an extracellular matrix-associated angiogenic inducer that functions as a ligand of integrin receptors to promote cell adhesion, migration, and proliferation. Aberrant expression of Cyr61 is associated with breast cancer, wound healing, and vascular diseases such as atherosclerosis and restenosis. To understand the functions of CYR61 during development, we have disrupted the Cyr61 gene in mice. We show here that Cyr61-null mice suffer embryonic death: approximately 30% succumbed to a failure in chorioallantoic fusion, and the reminder perished due to placental vascular insufficiency and compromised vessel integrity. These findings establish CYR61 as a novel and essential regulator of vascular development. CYR61 deficiency results in a specific defect in vessel bifurcation (nonsprouting angiogenesis) at the chorioallantoic junction, leading to an undervascularization of the placenta without affecting differentiation of the labyrinthine syncytiotrophoblasts. This unique phenotype is correlated with impaired Vegf-C expression in the allantoic mesoderm, suggesting that CYR61-regulated expression of Vegf-C plays a role in vessel bifurcation. The genetic and molecular basis of vessel bifurcation is presently unknown, and these findings provide new insight into this aspect of angiogenesis.

CYR61 (CCN1) is an extracellular matrix-associated protein of the CCN family, which also includes CTGF (CCN2), NOV (CCN3), WISP-1 (CCN4), WISP-2 (CCN5), and WISP-3 (CCN6). Purified CYR61 induces neovascularization in corneal implants, and Cyr61-null mice suffer embryonic death due to vascular defects, thus establishing that CYR61 is an important regulator of angiogenesis. Aberrant expression of Cyr61 is associated with breast cancer, wound healing, and vascular diseases such as atherosclerosis and restenosis. In culture, CYR61 functions through integrin-mediated pathways to promote cell adhesion, migration, and proliferation. Here we show that CYR61 can also promote cell survival and tubule formation in human umbilical vein endothelial cells. Furthermore, we have dissected the integrin receptor requirements of CYR61 with respect to its pro-angiogenic activities. Thus, CYR61-induced cell adhesion and tubule formation occur through interaction with integrin alpha(6)beta(1) in early passage endothelial cells in which integrins have not been activated. By contrast, in endothelial cells in which integrins are activated by phorbol ester or vascular endothelial growth factor, CYR61-promoted cell adhesion, migration, survival, growth factor-induced mitogenesis, and endothelial tubule formation are all mediated through integrin alpha(v)beta(3). These findings indicate that CYR61 is an activation-dependent ligand of integrin alpha(v)beta(3) and an activation-independent ligand of integrin alpha(6)beta(1) and that these integrins differentially mediate the pro-angiogenic activities of CYR61. These findings help to define the mechanisms by which CYR61 acts as an angiogenic regulator, provide a molecular interpretation for the loss of vascular integrity and increased apoptosis of vascular cells in Cyr61-null mice, and underscore the importance of CYR61 in the development and homeostasis of the vascular system.

A proposal is put forth to unify the nomenclature of the CCN family of secreted, cysteine rich regulatory proteins. In the order of their description in the literature, CCN1 (CYR61), CCN2 (CTGF), CCN3 (NOV), CCN4 (WISP-1), CCN5 (WISP-2), and CCN6 (WISP-3) constitute a family of matricellular proteins that regulate cell adhesion, migration, proliferation, survival, and differentiation, at least in part through integrin mediated mechanisms. This proposal is endorsed by the International CCN Society and will serve to eliminate confusion from the multiple names that have been given to these molecules.

CCN3 (NOV) is a matricellular protein of the CCN family, which also includes CCN1 (CYR61), CCN2 (CTGF), CCN4 (WISP-1), CCN5 (WISP-2), and CCN6 (WISP-3). During development, CCN3 is expressed widely in derivatives of all three germ layers, and high levels of expression are observed in smooth muscle cells of the arterial vessel wall. Altered expression of CCN3 has been observed in a variety of tumors, including hepatocellular carcinomas, Wilm's tumors, Ewing's sarcomas, gliomas, rhabdomyosarcomas, and adrenocortical carcinomas. To understand its biological functions, we have investigated the activities of purified recombinant CCN3. We show that in endothelial cells, CCN3 supports cell adhesion, induces directed cell migration (chemotaxis), and promotes cell survival. Mechanistically, CCN3 supports human umbilical vein endothelial cell adhesion through multiple cell surface receptors, including integrins alphavbeta3, alpha5beta1, alpha6beta1, and heparan sulfate proteoglycans. In contrast, CCN3-induced cell migration is dependent on integrins alphavbeta3 and alpha5beta1, whereas alpha6beta1 does not play a role in this process. Although CCN3 does not contain a RGD sequence, it binds directly to immobilized integrins alphavbeta3 and alpha5beta1, with half-maximal binding occurring at 10 nm and 50 nm CCN3, respectively. Furthermore, CCN3 induces neovascularization when implanted in rat cornea, demonstrating that it is a novel angiogenic inducer. Together, these findings show that CCN3 is a ligand of integrins alphavbeta3 and alpha5beta1, acts directly upon endothelial cells to stimulate pro-angiogenic activities, and induces angiogenesis in vivo.

Wnt-induced secreted protein 1 (WISP-1/CCN4) is a member of the CCN family that is highly expressed in skeletal tissue and in osteoprogenitor cells induced to differentiate in vitro. To determine the function of WISP-1 during osteogeneis, osteogenic bone marrow stromal cells (BMSCs) were transduced with WISP-1 adenovirus (adWISP-1) in the presence or absence of bone morphogenetic protein 2 (BMP-2) adenovirus (adBMP-2). WISP-1 overexpression enhanced the ability of BMP-2 to direct BMSCs toward osteogenic differentiation and appeared to work by stimulating Smad-1/5/8 phosphorylation and activation. The ability of WISP-1 to enhance BMP-2 activity also was shown in vivo using an ectopic osteogenesis assay with BMSCs transduced with WISP-1, BMP-2, or both. When BMSCs were infected with lentivirus containing human WISP1 shRNA, they formed less bone in vivo and were less responsive to BMP-2, confirming that WISP-1 and BMP-2 have a functional interaction. Immunoprecipitation (IP) and Western blot analysis showed that WISP-1 bound directly to BMP-2 and showed that WISP-1 increased BMP-2 binding to hBMSCs in a dose-dependent fashion. To understand how WISP-1 enhanced BMP-2 signaling, the influence of WISP-1 on integrin expression was analyzed. WISP-1 induced the mRNA and protein levels of α(5)-integrin and, further, was found to bind to it. Antibody-blocking experiments showed that the BMP-2 binding to BMSCs that was enhanced by WISP-1 was completely neutralized by treatment with anti-integrin α(5)β(1) antibody. Pilot studies and the use of transgenic mice that overexpressed human WISP-1 in preosteoblasts had increased bone mineral density (BMD), trabecular thickness, and bone volume (BV/TV) over wild-type controls, supporting observations using human osteoprogenitors that WISP-1 has a positive influence on osteogenesis in vivo. In conclusion, these studies show, for the first time, that WISP-1 has a positive influence on bone cell differentiation and function and may work by enhancing the effects of BMP-2 to increase osteogenesis through a mechanism potentially involving binding to integrin α(5)β(1).

The novel angiogenic inducer CCN3 (NOV, nephroblastoma overexpressed) is a matricellular protein of the CCN family, which also includes CCN1 (CYR61), CCN2 (CTGF), CCN4 (WISP-1), CCN5 (WISP-2), and CCN6 (WISP-3). CCN3 is broadly expressed in derivatives of all three germ layers during mammalian development, and its deranged expression is associated with vascular injury and a broad range of tumors. We have shown that CCN3 promotes proangiogenic activities in vascular endothelial cells through integrin receptors and induces neovascularization in vivo (Lin, C. G., Leu, S. J., Chen, N., Tebeau, C. M., Lin, S. X., Yeung, C. Y., and Lau, L. F. (2003) J. Biol. Chem. 278, 24200-24208). In this study, we show that CCN3 is highly expressed in granulation tissue of cutaneous wounds 5-7 days after injury and is capable of inducing responses in primary fibroblasts consistent with wound healing. Purified CCN3 supports primary skin fibroblast adhesion through integrins alpha(5)beta(1) and alpha(6)beta(1) and induces fibroblast chemotaxis through integrin alpha(v)beta(5). We show that CCN3 is a novel ligand of alpha(v)beta(5) in a solid phase binding assay. Although not mitogenic on its own, CCN3 also enhances basic fibroblast growth factor-induced DNA synthesis. Furthermore, CCN3 up-regulates MMP-1 and PAI-1 expression but interacts with TGF-beta1 in an antagonistic or synergistic manner to regulate the expression of specific genes. These findings, together with its angiogenic activity, support a role for CCN3 in cutaneous wound healing in skin fibroblasts and establish its matricellular mode of action through integrin receptors.

The CCN family of genes constitutes six members of small secreted cysteine rich proteins, which exists only in vertebrates. The major members of CCN are CCN1 (Cyr61), CCN2 (CTGF), and CCN3 (Nov). CCN4, CCN5, and CCN6 were formerly reported to be in the Wisp family, but they are now integrated into CCN due to the resemblance of their four principal modules: insulin like growth factor binding protein, von Willebrand factor type C, thrombospondin type 1, and carboxy-terminal domain. CCNs show a wide and highly variable expression pattern in adult and in embryonic tissues, but most studies have focused on their principal role in osteo/chondrogenesis and vasculo/angiogenesis from the aspect of migration, growth, and differentiation of mesenchymal cells. CCN proteins simultaneously integrate and modulate the signals of integrins, bone morphogenetic protein, vascular endothelial growth factor, Wnt, and Notch by direct binding. However, the priority in the use of the signals is different depending on the cell status. Even the equivalent counterparts show a difference in signal usage among species. It may be that the evolution of the CCN family continues to keep pace with vertebrate evolution itself.

Chondrogenesis is a process involving stem-cell differentiation through the coordinated effects of growth/differentiation factors and extracellular matrix (ECM) components. Recently, mesenchymal stem cells (MSCs) were found within the cartilage, which constitutes a specific niche composed of ECM proteins with unique features. Therefore, we hypothesized that the induction of MSC differentiation towards chondrocytes might be induced and/or influenced by molecules from the microenvironment. Using microarray analysis, we previously identified genes that are regulated during MSC differentiation towards chondrocytes. In this study, we wanted to precisely assess the differential expression of genes associated with the microenvironment using a large-scale real-time PCR assay, according to the simultaneous detection of up to 384 mRNAs in one sample. Chondrogenesis of bone-marrow-derived human MSCs was induced by culture in micropellet for various periods of time. Total RNA was extracted and submitted to quantitative RT-PCR. We identified molecules already known to be involved in attachment and cell migration, including syndecans, glypicans, gelsolin, decorin, fibronectin, and type II, IX and XI collagens. Importantly, we detected the expression of molecules that were not previously associated with MSCs or chondrocytes, namely metalloproteases (MMP-7 and MMP-28), molecules of the connective tissue growth factor (CTGF); cef10/cyr61 and nov (CCN) family (CCN3 and CCN4), chemokines and their receptors chemokine CXC motif ligand (CXCL1), Fms-related tyrosine kinase 3 ligand (FlT3L), chemokine CC motif receptor (CCR3 and CCR4), molecules with A Disintegrin And Metalloproteinase domain (ADAM8, ADAM9, ADAM19, ADAM23, A Disintegrin And Metalloproteinase with thrombospondin type 1 motif ADAMTS-4 and ADAMTS-5), cadherins (4 and 13) and integrins (alpha4, alpha7 and beta5). Our data suggest that crosstalk between ECM components of the microenvironment and MSCs within the cartilage is responsible for the differentiation of MSCs into chondrocytes.

The CCN family of genes currently comprises six secreted proteins (designated CCN1-6 after Cyr61/CCN1; ctgf/CCN2; Nov/CCN3; WISP1/CCN4; WISP2/CCN5, WISP3/CCN6) with a similar mosaic primary structure. It is now well accepted that CCN proteins are not growth factors but matricellular proteins that modify signaling of other molecules, in particular those associated with the extracellular matrix. CCN proteins are involved in mitosis, adhesion, apoptosis, extracellular matrix production, growth arrest and migration of multiple cell types. Since their first identification as matricellular factors, the CCN proteins now figure prominently in a variety of major diseases and are now considered valid candidates for therapeutic targeting. Dissection of the molecular mechanisms governing the biological properties of these proteins is being actively pursued by an expanding network of scientists around the globe who will meet this year at the 5th International Workshop on the CCN family of Genes, organized by the International CCN Society ( http://ccnsociety.com ), home for an international cadre of collaborators working in the CCN field.

The CCN proteins contain six members, namely CCN1 to CCN6, which are small secreted cysteine-rich proteins. The CCN proteins are modular proteins, containing up to four functional domains. Many of the CCN members are induced by growth factors, cytokines, or cellular stress. The CCNs show a wide and highly variable expression pattern in adult and in embryonic tissues. The CCN proteins can integrate and modulate the signals of integrins, BMPs, VEGF, Wnts, and Notch. The involvement of integrins in mediating CCN signaling may provide diverse context-dependent responses in distinct cell types. CCN1 and CCN2 play an important role in development, angiogenesis and cell adhesion, whereas CCN3 is critical to skeletal and cardiac development. CCN4, CCN5 and CCN6 usually inhibit cell growth. Mutations of Ccn6 are associated with the progressive pseudorheumatoid dysplasia and spondyloepiphyseal dysplasia tarda. In stem cell differentiation, CCN1, CCN2, and CCN3 play a principal role in osteogenesis, chondrogenesis, and angiogenesis. Elevated expression of CCN1 is associated with more aggressive phenotypes of human cancer, while the roles of CCN2 and CCN3 in tumorigenesis are tumor type-dependent. CCN4, CCN5 and CCN6 function as tumor suppressors. Although CCN proteins may play important roles in fine-tuning other major signaling pathways, the precise function and mechanism of action of these proteins remain undefined. Understanding of the biological functions of the CCN proteins would not only provide insight into their roles in numerous cellular processes but also offer opportunities for developing therapeutics by targeting CCN functions.

The CCN family of proteins includes six members presently known as CCN1, CCN2, CCN3, CCN4, CCN5 and CCN6. These proteins were originally designated CYR61, CTGF, NOV, and WISP-1, WISP-2, WISP-3. Although these proteins share a significant amount of structural features and a partial identity with other large families of regulatory proteins, they exhibit different biological functions. A critical examination of the progress made over the past two decades, since the first CCN proteins were discovered brings me to the conclusion that most of our present knowledge regarding the functions of these proteins was predicted very early after their discovery. In an effort to point out some of the gaps that prevent us to reach a comprehensive view of the functional interactions between CCN proteins, it is necessary to reconsider carefully data that was already published and put aside, either because the scientific community was not ready to accept them, or because they were not fitting with the « consensus » when they were published. This review article points to avenues that were not attracting the attention that they deserved. However, it is quite obvious that the six members of this unique family of tetra-modular proteins must act in concert, either simultaneously or sequentially, on the same sites or at different times in the life of living organisms. A better understanding of the spatio-temporal regulation of CCN proteins expression requires considering the family as such, not as a set of single proteins related only by their name. As proposed in this review, there is enough convincing pieces of evidence, at the present time, in favor of these proteins playing a role in the coordination of multiple signaling pathways, and constituting a Centralized Communication Network. Deciphering the hierarchy of regulatory circuits involved in this complex system is an important challenge for the near future. In this article, I would like to briefly review the concept of a CCN family of proteins and critically examine the progress made over the past 10 years in the understanding of their biological functions and involvement in both normal and pathological processes.

Wnt1 inducible signaling pathway protein 1 (WISP1/CCN4) is a CCN family member more broadly identified with development and tumorigenesis. However, recent studies have shed new light and enthusiasm on WISP1 as a novel target directed against toxic cell degeneration. Here we show WISP1 prevents apoptotic degeneration in primary neurons during oxidant stress through the activation of protein kinase B (Akt1), the post-translational maintenance of β-catenin integrity that is consistent with inhibition of glycogen synthase kinase-3β (GSK-3β), and the subcellular trafficking of β- catenin to foster its translocation to the nucleus. Interestingly, WISP1 autoregulates its expression through the promotion of β-catenin activity and may employ β-catenin to have a limited control over autophagy, but neuronal injury during oxidant stress as a result of autophagy appears portioned to a small population of neurons without significant impact upon overall cell survival. New strategies that target WISP1, its autoregulation, and the pathways responsible for neuronal cell injury may bring forth new insight for the treatment of neurodegenerative disorders.

Prostate cancer (PC) is a leading cause of death in men however the factors that regulate its progression and eventual metastasis to bone remain unclear. Here we show that WISP1/CCN4 expression in prostate cancer tissues was up-regulated in early stages of the disease and, further, that it correlated with increased circulating levels of WISP1 in the sera of patients at early stages of the disease. WISP1 was also elevated in the mouse prostate cancer model TRAMP in the hypoplastic diseased tissue that develops prior to advanced carcinoma formation. When the ability of anti-WISP1 antibodies to reduce the spread of PC3-Luc cells to distant sites was tested it showed that twice weekly injections of anti-WISP1 antibodies reduced the number and overall size of distant tumors developed after intracardiac (IC) injection of PC3-Luc cells in mice. The ability of antibodies against WISP1 to inhibit growth of PC3-Luc cancer cells in mice was also evaluated and showed that twice weekly injections of anti-WISP1 antibodies reduced local tumor growth when examined in xenografts. To better understand the mechanism of action, the migration of PC3-Luc cells through membranes with or without a Matrigel™ barrier showed the cells were attracted to WISP1, and that this attraction was inhibited by treatment with anti-WISP1 antibodies. We also show the expression of WISP1 at the bone-tumor interface and in the stroma of early grade cancers suggested WISP1 expression is well placed to play roles in both fostering growth of the cancer and its spread to bone. In summary, the up-regulation of WISP1 in the early stages of cancer development coupled with its ability to inhibit spread and growth of prostate cancer cells makes it both a potential target and an accessible diagnostic marker for prostate cancer.

OBJECTIVE

Referred to as CCN, the family of growth factors consisting of cystein-rich protein 61 (CYR61, also known as CCN1), connective tissue growth factor (CTGF, also known as CCN2), nephroblastoma overexpressed gene (NOV, also known as CCN3) and WNT1-inducible signalling pathway proteins 1, 2 and 3 (WISP1, -2 and -3; also known as CCN4, -5 and -6) affects cellular growth, differentiation, adhesion and locomotion in wound repair, fibrotic disorders, inflammation and angiogenesis. AGEs formed in the diabetic milieu affect the same processes, leading to diabetic complications including diabetic retinopathy. We hypothesised that pathological effects of AGEs in the diabetic retina are a consequence of AGE-induced alterations in CCN family expression.

METHODS

CCN gene expression levels were studied at the mRNA and protein level in retinas of control and diabetic rats using real-time quantitative PCR, western blotting and immunohistochemistry at 6 and 12 weeks of streptozotocin-induced diabetes in the presence or absence of aminoguanidine, an AGE inhibitor. In addition, C57BL/6 mice were repeatedly injected with exogenously formed AGE to establish whether AGE modulate retinal CCN growth factors in vivo.

RESULTS

After 6 weeks of diabetes, Cyr61 expression levels were increased more than threefold. At 12 weeks of diabetes, Ctgf expression levels were increased twofold. Treatment with aminoguanidine inhibited Cyr61 and Ctgf expression in diabetic rats, with reductions of 31 and 36%, respectively, compared with untreated animals. Western blotting showed a twofold increase in CTGF production, which was prevented by aminoguanidine treatment. In mice infused with exogenous AGE, Cyr61 expression increased fourfold and Ctgf expression increased twofold in the retina.

CONCLUSIONS

CTGF and CYR61 are downstream effectors of AGE in the diabetic retina, implicating them as possible targets for future intervention strategies against the development of diabetic retinopathy.

The CCN family of proteins comprises the members CCN1, CCN2, CCN3, CCN4, CCN5 and CCN6. They share four evolutionarily conserved functional domains, and usually interact with various cytokines to elicit different biological functions including cell proliferation, adhesion, invasion, migration, embryonic development, angiogenesis, wound healing, fibrosis and inflammation through a variety of signalling pathways. In the past two decades, emerging functions for the CCN proteins (CCNs) have been identified in various types of cancer. Perturbed expression of CCNs has been observed in a variety of malignancies. The aberrant expression of certain CCNs is associated with disease progression and poor prognosis. Insight into the detailed mechanisms involved in CCN-mediated regulation may be useful in understanding their roles and functions in tumorigenesis and cancer metastasis. In this review, we briefly introduced the functions of CCNs, especially in cancer.

OBJECTIVE

The expression of Wingless/int-1 (Wnt)-induced secreted protein-1 (WISP-1/CCN4), a member of the Cyr61-CTGF-Nov (CCN) family, has been examined in several types of cancer. However, the correlation between the WISP-1 expression and the clinical features of esophageal squamous cell carcinoma (ESCC) remain to be elucidated. This study aimed to clarify the expression of WISP-1 protein in patients with ESCC and also to examine the function of WISP-1 in esophageal cancer cells in vitro.

METHODS

One-hundred and ninety patients with thoracic esophageal carcinoma underwent transthoracic subtotal esophagectomy-between 2005 and 2009. All patients that had received previous therapy were excluded and 105 out of the 190 ESCC samples were analyzed immunohistochemically using WISP-1 antibody. The expression of WISP-1 mRNA in esophageal cancer cell lines was analyzed by RT-PCR. Growth assay and invasion assays were performed using WISP-1 transfected cells.

RESULTS

The immunohistochemical analysis showed that WISP-1-positive cases were closely associated with tumor size, tumor type, lymph node metastasis and poor prognosis. There were significantly more WISP-1-positive infiltrative type tumors than expanding type tumors. In the esophageal cancer cell lines examined, only TE8 expressed WISP-1 mRNA. The growth of WISP-1-transfected cells was significantly increased in comparison to the control cells, but no differences in the invasion activity were observed between the transfected cells and control cells.

CONCLUSIONS

The expression of WISP-1 may play an important role in the progression of ESCC. WISP-1 could therefore be a clinical marker for a poor prognosis in patients with ESCC and may also be a therapeutic target to control the progression of ESCC.

More than 110 million individuals will suffer from cognitive loss worldwide by the year 2050 with a majority of individuals presenting with Alzheimer's disease (AD). Yet, successful treatments for etiologies that involve β.-amyloid (Aβ.) toxicity in AD remain elusive and await novel avenues for drug development. Here we show that Wnt1 inducible signaling pathway protein 1 (WISP1/CCN4) controls the post-translational phosphorylation of Akt1, p70S6K, and AMP activated protein kinase (AMPK) to the extent that tuberous sclerosis complex 2 (TSC2) (Ser1387) phosphorylation, a target of AMPK, is decreased and TSC2 (Thr1462) phosphorylation, a target of Akt1, is increased. The ability of WISP1 to limit TSC2 activity allows WISP1 to increase the activity of p70S6K, since gene silencing of TSC2 further enhances WISP1 phosphorylation of p70S6K. However, a minimal level of TSC2 activity is necessary to modulate WISP1 cytoprotection that may require modulation of mTOR activity, since gene knockdown of TSC2 impairs the ability of WISP1 to protect microglia against apoptotic membrane phosphatidylserine (PS) exposure, nuclear DNA degradation, mitochondrial membrane depolarization, and cytochrome c release during Aβ. exposure.

Given the present challenges to attain effective treatment for β-amyloid (Aβ) toxicity in neurodegenerative disorders such as Alzheimer's disease, development of novel cytoprotective pathways that can assist immune mediated therapies through the preservation of central nervous system microglia could offer significant promise. We show that the CCN4 protein, Wnt1 inducible signaling pathway protein 1 (WISP1), is initially up-regulated by Aβ and can modulate its endogenous expression for the protection of microglia during Aβ mediated apoptosis. WISP1 activates mTOR and phosphorylates p70S6K and 4EBP1 through the control of the regulatory mTOR component PRAS40. Loss of PRAS40 through gene reduction or inhibition by WISP1 is cytoprotective. WISP1 ultimately governs PRAS40 by sequestering PRAS40 intracellularly through post-translational phosphorylation and binding to protein 14-3-3. Our work identifies WISP1, mTOR signaling, and PRAS40 as targets for new strategies directed against Alzheimer's disease and related disorders.

As a member of the secreted extracellular matrix associated proteins of the CCN family, Wnt1 inducible signaling pathway protein 1 (WISP1/CCN4) is garnering increased attention not only as a potent proliferative entity, but also as a robust cytoprotective agent during toxic insults. Here we demonstrate that WISP1 prevents forkhead transcription factor FoxO3a mediated caspase 1 and caspase 3 apoptotic cell death in primary neurons during oxidant stress. Phosphoinositide 3-kinase (PI 3-K) and protein kinase B (Akt1) are necessary for WISP1 to foster posttranslational phosphorylation of FoxO3a and sequester FoxO3a in the cytoplasm of neurons with protein 14-3-3. Through an autoregulatory loop, WISP1 also minimizes deacytelation of FoxO3a, prevents caspase 1 and 3 activation, and promotes an effective neuroprotective level of SIRT1 activity through SIRT1 nuclear trafficking and prevention of SIRT1 caspase degradation. Elucidation of the critical pathways of WISP1 that determine neuronal cell survival during oxidative stress may offer novel therapeutic avenues for neurodegenerative disorders.

Diabetes mellitus affects almost 350 million individuals throughout the globe resulting in significant morbidity and mortality. Of further concern is the growing population of individuals that remain undiagnosed but are susceptible to the detrimental outcomes of this disorder. Diabetes mellitus leads to multiple complications in the central and peripheral nervous systems that include cognitive impairment, retinal disease, neuropsychiatric disease, cerebral ischemia, and peripheral nerve degeneration. Although multiple strategies are being considered, novel targeting of trophic factors, Wnt signaling, Wnt1 inducible signaling pathway protein 1, and stem cell tissue regeneration are considered to be exciting prospects to overcome the cellular mechanisms that lead to neuronal injury in diabetes mellitus involving oxidative stress, apoptosis, and autophagy. Pathways that involve insulin-like growth factor-1, fibroblast growth factor, epidermal growth factor, and erythropoietin can govern glucose homeostasis and are intimately tied to Wnt signaling that involves Wnt1 and Wnt1 inducible signaling pathway protein 1 (CCN4) to foster control over stem cell proliferation, wound repair, cognitive decline, β-cell proliferation, vascular regeneration, and programmed cell death. Ultimately, cellular metabolism through Wnt signaling is driven by primary metabolic pathways of the mechanistic target of rapamycin and AMP activated protein kinase. These pathways offer precise biological control of cellular metabolism, but are exquisitely sensitive to the different components of Wnt signaling. As a result, unexpected clinical outcomes can ensue and therefore demand careful translation of the mechanisms that govern neural repair and regeneration in diabetes mellitus.

The CCN family of proteins (CCN1, CCN2, CCN3, CCN4, CCN5 and CCN6) are multifunctional mosaic proteins that play keys roles in crucial areas of physiology such as angiogenesis, skeletal development tumourigenesis, cell proliferation, adhesion and survival. This expansive repertoire of functions comes through a modular structure of 4 discrete domains that act both independently and in concert. How these interactions with ligands and with neighbouring domains lead to the biological effects is still to be explored but the molecular structure of the domains is likely to play an important role in this. In this review we have highlighted some of the key features of the individual domains of CCN family of proteins based on their biological effects using a homology modelling approach.

CCN4/Wnt-induced secreted protein 1 (WISP1) is one of the CCN (CTGF/Cyr61/Nov) family proteins. CCN members have typical structures composed of four conserved cysteine-rich modules and their variants lacking certain modules, generated by alternative splicing or gene mutations, have been described in various pathological conditions. Several previous reports described a CCN4/WISP1 variant (WISP1v) lacking the second module in a few malignancies, but no information concerning the production of WISP1 variants in normal tissue is currently available. The expression of CCN4/WISP1 mRNA and its variants were analyzed in a human chondrosarcoma-derived chondrocytic cell line, HCS-2/8, and primary rabbit growth cartilage (RGC) chondrocytes. First, we found WISP1v and a novel variant of WISP1 (WISP1vx) to be expressed in HCS-2/8, as well as full-length WISP1 mRNA. This new variant was lacking the coding regions for the second and third modules and a small part of the first module. To monitor the expression of CCN4/WISP1 mRNA along chondrocyte differentiation, RGC cells were cultured and sampled until they were mineralized. As a result, we identified a WISP1v ortholog in normal RGC cells. Interestingly, the WISP1v mRNA level increased dramatically along with terminal differentiation. Furthermore, overexpression of WISP1v provoked expression of an alkaline phosphatase gene that is a marker of terminal differentiation in HCS-2/8 cells. These findings indicate that WISP1v thus plays a critical role in chondrocyte differentiation toward endochondral ossification, whereas HCS-2/8-specific WISP1vx may be associated with the transformed phenotypes of chondrosarcomas.

WISP1 (Wnt1-inducible signaling pathway protein-1, also known as CCN4) is a member of the secreted extracellular matrix-associated proteins of the CCN family and a target gene of the Wingless-type (WNT) signaling pathway. Growing evidence links the WNT signaling pathway to the regulation of adipogenesis and low-grade inflammation in obesity. We aimed to validate WISP1 as a novel adipokine. Human adipocyte differentiation was associated with increased WISP1 expression and secretion. Stimulation of human macrophages with WISP1 led to a proinflammatory response. Circulating WISP1 and WISP1 subcutaneous adipose tissue expression were regulated by weight changes in humans and mice. WISP1 expression in visceral and subcutaneous fat tissue was associated with markers of insulin resistance and inflammation in glucose-tolerant subjects. In patients with nonalcoholic fatty liver disease, we found no correlation among disease activity score, liver fat content, and WISP1 expression. Insulin regulated WISP1 expression in adipocytes in vitro but had no acute effect on WISP1 gene expression in subcutaneous fat tissue in overweight subjects who had undergone hyperinsulinemic clamp experiments. The data suggest that WISP1 may play a role in linking obesity to inflammation and insulin resistance and could be a novel therapeutic target for obesity.