Venous leg ulcers: patient concordance with compression therapy and its impact on healing and prevention of recurrence.

Moffatt Christine, Kommala Dheerendra, Dourdin Nathalie, Choe Yoonhee. Venous leg ulcers: patient concordance with compression therapy and its impact on healing and prevention of recurrence. ABSTRACT This study aimed to review available data on the reasons attributed to patient non concordance with compression therapy for the treatment of venous leg ulcers (VLUs), the frequency of non concordance and its effects on clinical outcomes. The biomedical literature was searched for publications on VLUs, compression therapy and concordance over the past 20 years. Physical, aesthetic and cosmetic factors, patient lack of education about VLUs, cost of therapy and issues with treatment by clinicians were all reported to influence concordance with compression therapy. The search identified 10 studies reporting patient concordance with compression stockings or bandages; while non concordance ranged from 2% to 42% of patients in three randomised controlled trials, it was generally higher in real-world studies, ranging from 9.7% to 80%. Another set of six studies indicated that the healing rate was half and the median time to complete healing was twice as long when patients were not concordant. Further, recurrence rates were 2-20 times greater when patients did not comply with the use of stockings following VLU healing. In conclusion, published biomedical literature has documented that non concordance with compression therapy negatively impacts the outcome of VLUs, highlighting the need to improve patient concordance to maximise therapeutic benefits.

Transactivation of capn2 by myogenic regulatory factors during myogenesis.

The calcium-activated cysteine protease m-calpain plays a pivotal role during the earlier stages of myogenesis, particularly during fusion. The enzyme is a heterodimer, encoded by the genes capn2, for the large subunit, and capn4, for the small subunit. To study the regulation of m-calpain, the DNA sequence upstream of capn2 was analyzed for promoter elements, revealing the existence of five consensus-binding sites (E-box) for several myogenic regulatory factors and one binding site for myocyte enhancer factor-2 (MEF-2). Transient transfections with reporter gene constructs containing the E-box revealed that MyoD presents a high level of transactivation of reporter constructs containing this region, in particular the sequences including the MEF-2/E4-box. In addition, over-expression of various myogenic factors demonstrated that MyoD and myogenin with much less efficiency, can up-regulate capn2, both singly and synergistically, while Myf5 has no effect on synthesis of the protease. Experiments with antisense oligonucleotides directed against each myogenic factor revealed that MyoD plays a specific and pivotal role during capn2 regulation, and cannot be replaced wholly by myogenin and Myf5.

PTEN deficiency in a luminal ErbB-2 mouse model results in dramatic acceleration of mammary tumorigenesis and metastasis.

Overexpression and/or amplification of the ErbB-2 oncogene as well as inactivation of the PTEN tumor suppressor are two important genetic events in human breast carcinogenesis. To address the biological impact of conditional inactivation of PTEN on ErbB-2-induced mammary tumorigenesis, we generated a novel transgenic mouse model that utilizes the murine mammary tumor virus (MMTV) promoter to directly couple expression of activated ErbB-2 and Cre recombinase to the same mammary epithelial cell (MMTV-NIC). Disruption of PTEN in the mammary epithelium of the MMTV-NIC model system dramatically accelerated the formation of multifocal and highly metastatic mammary tumors, which exhibited homogenous pathology. PTEN-deficient/NIC-induced tumorigenesis was associated with an increase in angiogenesis. Moreover, inactivation of PTEN in the MMTV-NIC mouse model resulted in hyperactivation of the phosphatidylinositol 3'-kinase/Akt signaling pathway. However, like the parental strain, tumors obtained from PTEN-deficient/NIC mice displayed histopathological and molecular features of the luminal subtype of primary human breast cancer. Taken together, our findings provide important implications in understanding the molecular determinants of mammary tumorigenesis driven by PTEN deficiency and ErbB-2 activation and could provide a valuable tool for testing the efficacy of therapeutic strategies that target these critical signaling pathways.

Phosphatase and tensin homologue deleted on chromosome 10 deficiency accelerates tumor induction in a mouse model of ErbB-2 mammary tumorigenesis.

Loss of the tumor suppressor phosphatase and tensin homologue deleted on chromosome 10 (PTEN) and amplification or elevated expression of ErbB-2 are both involved in human breast cancer. To directly test the importance of these genetic events in mammary tumorigenesis, we have assessed whether mammary-specific disruption of PTEN could cooperate with activation of ErbB-2. Transgenic mice expressing ErbB-2 under the transcriptional control of its endogenous promoter (ErbB-2(KI)) were interbred with mice carrying conditional PTEN alleles and an MMTV/Cre transgene. Loss of one or both PTEN alleles resulted in a dramatic acceleration of mammary tumor onset and an increased occurrence of lung metastases in the ErbB-2(KI) strain. Tumor progression in PTEN-deficient/ErbB-2(KI) strains was associated with elevated ErbB-2 protein levels, which were not due to ErbB-2 amplification or to a dramatic increase in ErbB-2 transcripts. Moreover, the PTEN-deficient/ErbB-2(KI)-derived mouse mammary tumors display striking morphologic heterogeneity in comparison with the homogeneous pathology of the ErbB-2(KI) parental strain. Therefore, inactivation of PTEN would not only have a dramatic effect on ErbB-2-induced mammary tumorigenesis but would also lead to the formation of mammary tumors that, in part, display pathologic and molecular features associated with the basal-like subtype of primary human breast cancer.

Conditional disruption of ubiquitous calpains in the mouse.

Ubiquitous mu- and m-calpain proteases are implicated in development and apoptosis. They are heterodimers consisting of 80-kDa catalytic subunits encoded by capn1 and capn2, respectively, and a common 28-kDa regulatory subunit encoded by capn4. The regulatory subunit is required to maintain stability and activity of mu- and m-calpains; thus, genetic disruption of capn4 was predicted to eliminate both calpain activities. Germline disruption of capn4 caused embryonic lethality, hampering the use of those mouse models to explore physiological calpain functions. Here we describe a loxP/cre conditional capn4 targeted mouse model that enables tissue-specific and temporal deletion of calpain activity. Disruption of the floxed capn4 gene using a ubiquitous cytomegalovirus promoter driven Cre recombinase transgene led to midgestation embryonic lethality. Fibroblasts from these embryos lacked detectable regulatory subunit expression, had reduced levels of the mu- and m-calpain catalytic subunits, and had no detectable mu- and m-calpain activities. These defects were corrected with a capn4-encoding lentivirus.

Ubiquitous calpains promote caspase-12 and JNK activation during endoplasmic reticulum stress-induced apoptosis.

Ubiquitously expressed mu- and m-calpain proteases are implicated in development and apoptosis. They consist of 80-kDa catalytic subunits encoded by the capn1 and capn2 genes, respectively, and a common 28-kDa regulatory subunit encoded by the capn4 gene. The regulatory subunit is required to maintain the stability and activity of mu- and m-calpains. Accordingly, genetic disruption of capn4 in the mouse eliminated both ubiquitous calpain activities. In embryonic fibroblasts derived from these mice, calpain deficiency correlated with resistance to endoplasmic reticulum (ER) stress-induced apoptosis, and this was directly related to a calpain requirement for activation of both caspase-12 and the ASK1-JNK cascade. This study provides compelling genetic evidence for calpain's role in caspase-12 activation at the ER, and reveals a novel role for the ubiquitous calpains in ER-stress induced apoptosis and JNK activation.

Calpain is required for MMP-2 and u-PA expression in SV40 large T-antigen-immortalized cells.

The absence of both mu- and m-calpain activity, caused by disruption of the capn4 gene in mice, retarded migration, and disrupted the cytoskeleton, both in primary capn4(-/-) embryonic fibroblasts (mEF) and in capn4(-/-) mEF immortalized with SV40 large T-antigen (TAg). These results are thought to reflect the role of calpain in integrin signaling to the cytoskeleton. The integrins are also involved, together with matrix metalloproteinases (MMP) and plasminogen activators (PA), in cellular invasion. This study therefore aimed to establish whether links exist between the calpain, MMP, and PA systems, using both primary and TAg-immortalized capn4(+/+) and capn4(-/-) embryonic fibroblasts. Both Matrigel invasion, and expression of MMP-2 and u-PA activities, correlated with calpain expression in TAg-containing cells, but not in primary cells. MMP-2 mRNA synthesis also correlated with calpain expression in the presence of TAg, but u-PA mRNA synthesis was not so correlated. The results suggest that calpain acquires new regulatory roles in the presence of TAg. Calpain is also required for v-Src-mediated transformation. It appears that calpain may have previously unsuspected roles in oncogenic transformation.

Calpain and myogenesis: development of a convenient cell culture model.

Previous studies have led us to hypothesize that m-calpain plays a pivotal role in myoblast fusion through its involvement in cell membrane and cytoskeleton component reorganization. To support this hypothesis, a convenient and simple myoblast culture model using frozen embryonic myoblasts was developed, which resolved a number of problems inherent to cell primary culture. Biological assays on cultured myoblasts using different media to define the characteristics of the fusion process were first conducted. Proteinase was detectable before the initiation of the fusion process and was closely correlated to the phenomenon of fusion under each culture condition studied. In addition, the study of calpastatin showed that the initiation of fusion does not require a decrease in the level of this endogenous inhibitor of calpains and also confirmed that calpastatin may be implicated in the determination of the end of fusion. On the other hand, analysis of the evolution of myogenic factors revealed that myogenins, MyoD and Myf5, increase very significantly during the formation of multinucleated myotubes. Moreover, the antisense technique against myogenin is capable of preventing the process of fusion by 50%, confirming the pivotal role of this factor in the early stages of differentiation. The possible role of myogenic regulator factors on m-calpain gene expression is discussed.