Elevated TGF-β1 Expression Research Articles

Effect of magnetic microbeads on sustained and targeted delivery of transforming growth factor-beta-1 for rotator cuff healing in a rat rotator cuff repair model

Structural failure is a well-established complication of rotator cuff repair procedures. To evaluate the effect of magnetic microbeads, designed for precise drug delivery via magnetic force, on sustained transforming growth factor-beta-1 (TGF-β1) release and rotator cuff healing in a rat rotator cuff repair model. TGF-β1 laden microbeads were prepared, and baseline in vitro experiments included the magnetization of the microbeads and TGF-β1 release tests. In an in vivo experiment using a rat rotator cuff repair model on both shoulders, 72 rats were randomly assigned to three groups (24 per group): group A, conventional repair; group B, repair with and simple TGF-β1 injection; and group C, repair with magnet insertion into the humeral head and TGF-β1 laden microbead injection. Delivery of TGF-β1 was evaluated at 1 and 7 days after the intervention using PCR, Western blot, and immunohistochemistry. At 6 weeks post-intervention, rotator cuff healing was assessed using biomechanical and histological analysis. The in vitro experiments confirmed the magnetization property of the microbeads and sustained delivery of TGF-β1 for up to 10 days. No difference in the TGF-β1 expression was found at day 1 in vivo. However, at day 7, group C exhibited a significantly elevated expression of TGF-β1 in both PCR and Western blot analyses compared to groups A and B (all P < 0.05). Immunohistochemical analysis revealed a higher expression of TGF-β1 at the repair site in group C on day 7. At 6 weeks, biomechanical analysis demonstrated a significantly higher ultimate failure load in group C than in groups A and B (P < 0.05) and greater stiffness than in group A (P = 0.045). In addition, histological analysis showed denser and more regular collagen fibers with complete continuity to the bone in group C than in groups A and B, a statistically significant difference according to the semi-quantitative scoring system (all P < 0.05). The use of the TGF-β1 laden magnetic microbeads demonstrated sustained delivery of TGF-β1 to the repair site, improving rotator cuff healing.

Expression of the Pro-Fibrotic Marker Periostin in a Mouse Model of Duchenne Muscular Dystrophy.

Duchenne muscular dystrophy (DMD) is characterised by fibrotic tissue deposition in skeletal muscle. We assessed the role of periostin in fibrosis using mdx mice, an established DMD murine model, for which we conducted a thorough examination of periostin expression over a year. RNA and protein levels in diaphragm (DIA) muscles were assessed and complemented by a detailed histological analysis at 5 months of age. In dystrophic DIAs, periostin (Postn) mRNA expression significantly exceeded that seen in wildtype controls at all timepoints analysed, with the highest expression at 5 months of age (p < 0.05). We found Postn to be more consistently highly expressed at the earlier timepoints compared to established markers of fibrosis like transforming growth factor-beta 1 (Tgf-β1) and connective tissue growth factor (Ctgf). Immunohistochemistry confirmed a significantly higher periostin protein expression in 5-month-old mdx mice compared to age-matched healthy controls (p < 0.01), coinciding with a significant fibrotic area percentage (p < 0.0001). RT-qPCR also indicated an elevated expression of Tgf-β1, Col1α1 (collagen type 1 alpha 1) and Ctgf in mdx DIAs compared to wild type controls (p < 0.05) at 8- and 12-month timepoints. Accordingly, immunoblot quantification demonstrated elevated periostin (3, 5 and 8 months, p < 0.01) and Tgf-β1 (8 and 12 months, p < 0.001) proteins in the mdx muscle. These findings collectively suggest that periostin expression is a valuable marker of fibrosis in this relevant model of DMD. They also suggest periostin as a potential contributor to fibrosis development, with an early onset of expression, thereby offering the potential for timely therapeutic intervention and its use as a biomarker in muscular dystrophies.

Immunomodulation by xylan and carrageenan-type polysaccharides from red seaweeds: Anti-inflammatory, wound healing, cytoprotective, and anticoagulant activities

The immunomodulatory properties of the polysaccharides (carrageenan, xylan) from Chondrus crispus (CC), Ahnfeltiopsis devoniensis (AD), Sarcodiotheca gaudichaudii (SG) and Palmaria palmata (PP) algal species were studied. Using RAW264.7 macrophages, we investigated the proliferation and migration capacity of different extracts along with their immunomodulatory activities, including nitric oxide (NO) production, phagocytosis, and secretion of pro-inflammatory cytokines. Polysaccharides from C. crispus and S. gaudichaudii effectively mitigated inflammation and improved scratch-wound healing. Polysaccharide fractions extracted under cold conditions (25 °C), including CC-1A, SG-1A and SG-1B stimulated cell proliferation, while fractions extracted under hot conditions (95 °C), including CC-3A, CC-2B and A. devoniensis (AD-3A), inhibited cell proliferation after 48 h. Furthermore, RAW264.7 cells treated with the fractions CC-3A, AD-1A, and SG-2A significantly reduced LPS-stimulated NO secretion over 24 h. Phagocytosis was significantly improved by treatment with C. crispus (CC-2B, CC-3B) and A. devoniensis (AD-3A) fractions. RAW264.7 cells treated with the CC-2A and SG-1A fractions showed elevated TGF-β1 expression without affecting TNF-α expression at 24 h. Polysaccharide fractions of A. devoniensis (ι/κ hybrid carrageenan; AD-2A, AD-3A) showed the highest anti-coagulation activity. CC-2A and SG-1A fractions enhanced various bioactivities, suggesting they are candidates for skin-health applications. The carrageenan fractions (CC-3A: λ-, μ-carrageenan, SG-2A: ν-, ι-carrageenan) tested herein showed great potential for developing anti-inflammatory and upscaled skin-health applications.

Evaluation of Transforming Growth Factor Beta-1 in the vein wall of males with varicocele.

This study aims to investigate potential differences in the presence of Transforming Growth Factor-Beta 1 (TGF-β1) between the vein walls of patients with varicocele and those of healthy individuals. The study comprised a total of 40 participants, divided into two groups. The control group (Group 1) consisted of 20 patients who underwent coronary bypass surgery, while the varicocele group (Group 2) included 20 patients scheduled for varicocelectomy. The cytoplasmic and nuclear staining patterns of TGF-β1 immunohistochemistry were assessed in tissue samples under light microscopy, identifying any differences in TGF-β1 presence between varicocele patient vein walls and normal (saphenous) veins. The varicocele group demonstrated lower nuclear and cytoplasmic TGF-β1 staining rates compared to the control group. After controlling for the independent factor of age, significantly lower nuclear and cytoplasmic staining was still observed in the varicocele group. This study is the first of its kind to compare TGF-β1 staining in the vein walls of varicocele patients and healthy individuals. Previous studies focusing on varicose veins reported elevated TGF-β1 expression. Contrarily, our study observed lower TGF-β1 expression in varicocele patient veins, marking a unique contribution to the field.

TGF-β1 and IL-17A comediate the protumor phenotype of neutrophils to regulate the epithelial-mesenchymal transition in oral squamous cell carcinoma.

The role of neutrophils in cancer has been the subject of intense research in recent years. One major theme that has emerged is that not all neutrophils are equal in the field of cancer. However, it remains unclear what induces the protumorigenic or antitumorigenic phenotype predominate in tumor. Therefore, this study aimed to investigate what factors induce which of these two phenotypes of neutrophil predominate in OSCC and to explore the role of neutrophil polarization on tumor. Immunofluorescence and immunohistochemistry staining were used to observe neutrophil infiltration and the expression of TGF-β1 and IL-17A in OSCC tissues. Recombinant human TGF-β1 and IL-17A were used to modulate neutrophil polarization. OSCC cell (SCC9 and SAS cell lines) migration, proliferation, invasion, stemness, and EMT were analyzed after treatment with conditioned medium from TGF-β1/IL-17A-activated neutrophils. The levels of neutrophil-associated markers in OSCC tissues and peripheral blood were examined by immunofluorescence staining and quantitative PCR. Our data showed neutrophil infiltration and elevated expression of TGF-β1 and IL-17A in OSCC tissues. The cooperative effect of TGF-β1 and IL-17A promoted neutrophils to take on a protumor phenotype in vitro. TGF-β1/IL-17A-activated neutrophils remarkably induced cell migration, proliferation, invasion, stemness, and EMT in OSCC cells. Additionally, OSCC patients showed increased expression of MMP9 and decreased expression of CCL3 in circulating neutrophils. TGF-β1 and IL-17A cooperated to augment the protumor functions of neutrophils, thereby promoting the progression of OSCC cells. In addition, the combination of neutrophil-associated markers may serve as a predictive method to screen for patients with OSCC.

Targeting the phosphoinositide 3-kinase/AKT pathways by small molecules and natural compounds as a therapeutic approach for breast cancer cells.

The phosphoinositide 3-kinase/AKT/mTOR (PI3K/AkT/mTOR) pathway plays a pivotal role in the uncontrolled growth, migration and development of human breast cancer. The elevated expression of TGF-β1 increases the PI3K/AkT/mTOR activity in human breast cancer tissue and potentially motivates tumor metastasis and resistance to chemotherapy. Here, we investigated whether treatment with PI3K/AkT/mTOR dual inhibitor NVP-BEZ235 alone or in combination with caffeic acid phenyl ester (CAPE) could prevent TGF-β1 effects on breast cancer cells. MCF-7 human breast cancer cells were exposed to TGF-β1 for 14days and then were treated with/without NVP-BEZ235 and/or CAPE. Cell viability, apoptosis, CXCR4 surface expression and mRNA levels of CXCR4 and TWIST-1 were analyzed in all treated groups. We found that treatment of human breast cancer cells with acombination of NVP-BEZ235 and CAPE increased induction of cellular death. Although flow cytometry analysis demonstrated that NVP-BEZ235 alone treatment reduced CXCR4 expression while increasing CXCR4 mRNA level; when NVP-BEZ235 was combined with CAPE, inhibition of CXCR4 surface expression and enhancement of CXCR4 mRNA expression was diminished. In addition, TWIST-1 mRNA expression was down regulated in samples treated with both NVP-BEZ235 and CAPE. These altogether signified that NVP-BEZ235 in combination with CAPE showed improved therapeutic efficacy in breast cancer cells by decreasing apoptotic resistance and reduction of CXCR4 and TWIST-1 expression at mRNA level could be one of mechanism of action.

Compound Astragalus and Salvia miltiorrhiza extract inhibits hepatocarcinogenesis via modulating TGF-β/TβR and Imp7/8.

Compound Astragalus and Salvia miltiorrhiza extract (CASE) is a Chinese herbal formula consisting of astragalosides, astragalus polysaccharide and salvianolic acids extracted from Astragalus membranaceus and Salvia miltiorhiza. Previous studies by our group have demonstrated that CASE effectively suppresses diethylinitrosamine (DEN)-induced hepatocellular carcinoma (HCC) in rats via modulating transforming growth factor β/Mothers against decapentaplegic (TGFβ/Smad) signaling. To further elucidate the mechanism of CASE, the effects of CASE on TGF-β1, the serine/threonine kinase receptors of TGF-β [TGF-β receptor type-I (TβRI) and TβRII] and karyopherins [Importin 7 (Imp7) and Imp8], which are crucial for TGF-β/Smad signaling in fibro-hepatocarcinogenesis, were assessed in the present study using in vivo (DEN-induced HCC in rats) and in vitro [TGF-β1-stimulated rat myofibroblasts (MFBs) and HepG2 cells] models of fibro-hepatocarcinogenesis. Hematoxylin and eosin staining revealed that CASE may suppress inflammatory reactions and fibrosis in HCC as well as increasing the differentiation of HCC cells. Positive TGF-β1 staining was increased in HCC nodule areas and in adjacent normal liver tissues in DEN-treated rats, while TβRI staining was increased only in normal adjacent liver tissues. The elevated expression of TGF-β1, TβRI and TβRII was suppressed by CASE. CASE treatment also reduced glutathione S-transferase P 1 and Imp7/8 protein expression in fibro-hepatocarcinogenesis. In vitro experiments confirmed that CASE was able to decrease the expression of TβRI and TβRII in TGF-β1-stimulated MFBs and HepG2 cells. These results indicate that the anti-HCC effect of CASE may be achieved by mediating TGF-β/TβR and Imp7/8 protein expression, suggesting that CASE has multiple targets in HCC treatment.

MicroRNA-483-5p Modulates the Expression of Cartilage-Related Genes in Human Chondrocytes through Down-Regulating TGF-β1 Expression.

Transforming growth factor-β1 (TGF-β1) has been reported to improve chondrocytes phenotype and function. The expression levels of microRNA-483-5p (miR-483-5p), a potential regulator of TGF-β signaling pathway, were elevated in chondrocytes of patients with osteoarthritis. In this study, we aimed to explore the role of miR-483-5p for the expression of cartilage-related genes in chondrocytes. Human chondrocytes were harvested from femoral condyle and tibial plateau of different donors (n = 10) following amputation due to sarcomas not involving the joint space. The expression levels of miR-483-5p and TGF-β1 mRNA were measured by qRT-PCR. Runt-related transcription factor 2 (RUNX2) is a transcription factor involved in chondrocyte differentiation, and type II collagen-degrading matrix metalloproteinase13 (MMP13) contributes to cartilage degradation. qRT-PCR was also used to measure the levels of RUNX2 and MMP-13 mRNAs, as well as type II collagen alpha 1 (COL2A1) and aggrecan mRNAs. COL2A1 and aggrecan are major cartilage extracellular matrix proteins that are essential for normal cartilage function. The expression levels of miR-483-5p were significantly increased in chondrocytes from Passage 0 to 2, whereas the expression levels of TGF-β1 mRNA were marginally decreased. Passage 1 chondrocytes were employed for following experiments. MiR-483-5p overexpression reduced TGF-β1 expression, while miR-483-5p knockdown dramatically elevated TGF-β1 expression both at mRNA and protein levels. Further, miR-483-5p overexpression significantly decreased the levels of COL2A1 and aggrecan mRNAs, and increased those of RUNX2 and MMP13 mRNAs, by down-regulating TGF-β1 expression. These findings suggest that modulating miR-483-5p expression may contribute to maintaining the cartilage tissues.

Panax notoginseng saponins administration modulates pro- /anti-inflammatory factor expression and improves neurologic outcome following permanent MCAO in rats.

Ischemic stroke, particularly permanent occlusion, accounts for the overwhelming majority of all strokes. In addition to the occlusion of arteries, the inflammatory response plays a pivotal role in the severity of the cerebral injury and its clinical prognosis. Here, panax notoginseng saponins (PNS) extracted from a traditional Chinese herbal medicine was administered following permanent middle cerebral artery occlusion (MCAO) in rats to explore the neuroprotective mechanisms against ischemic injury. The results showed that MCAO surgery was successful in producing an infarct and that PNS and nimodipine could ameliorate the neurological deficits. The expression levels of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) and transforming growth factor-β1 (TGF-β1) were increased, while the level of interleukin-10 (IL-10) was reduced in the infarct cortex 7days after MCAO, as assessed by immunohistochemistry, western blotting and quantitative real-time PCR (qRT-PCR). PNS was able to markedly reduce the overexpression of IL-1β and TNF-α while significantly promoting the expression of IL-10, but did not affect the elevated expression of TGF-β1. Meanwhile, nimodipine was able to significantly reduce the expression of IL-1β and TNF-α, but had no obvious effect on IL-10 or TGF-β1. In addition, the serum levels of TNF-α, IL-10 and TGF-β1 were basically consistent with cerebral tissue results; however, the IL-1β levels did not differ. We conclude that PNS can directly down-regulate the overexpression of proinflammatory factors IL-1β and TNF-α while up-regulating the expression of anti-inflammatory factor IL-10 in the core region of the cerebral infarct, thereby preventing neurological damage in rats after permanent MCAO.

EIF5A2 predicts outcome in localised invasive bladder cancer and promotes bladder cancer cell aggressiveness in vitro and in vivo

Background:EIF5A2, eukaryotic translation initiation factor 5A2, is associated with several human cancers. In this study, we investigated the role of EIF5A2 in the metastatic potential of localised invasive bladder cancer (BC) and its underlying molecular mechanisms were explored.Methods:The expression pattern of EIF5A2 in localised invasive BC was determined by immunohistochemistry. In addition, the function of EIF5A2 in BC and its underlying mechanisms were elucidated with a series of in vitro and in vivo assays.Results:Overexpression of EIF5A2 was an independent predictor for poor metastasis-free survival of localised invasive BC patients treated with radical cystectomy. Knockdown of EIF5A2 inhibited BC cell migratory and invasive capacities in vitro and metastatic potential in vivo and reversed epithelial–mesenchymal transition (EMT), whereas overexpression of EIF5A2 promoted BC cells motility and invasiveness in vitro and metastatic potential in vivo and induced EMT. In addition, we found that EIF5A2 might activate TGF-β1 expression to induce EMT and drive aggressiveness in BC cells. EIF5A2 stabilized STAT3 and stimulated nuclear localisation of STAT3, which resulted in increasing enrichment of STAT3 onto TGF-β1 promoter to enhance the transcription of TGF-β1.Conclusions:EIF5A2 overexpression predicts tumour metastatic potential in patients with localised invasive BC treated with radical cystectomy. Furthermore, EIF5A2 elevated TGF-β1 expression through STAT3 to induce EMT and promotes aggressiveness in BC.

Expression of MMP-2, TIMP-2, TGF-β1, and Decorin in Dupuytren’s Contracture

To investigate the mechanisms underlying matrix deposition in Dupuytren’s disease, the expression of gelatinase A (MMP-2), the tissue inhibitor of metalloproteinase-2 (TIMP-2), transforming growth factor beta 1 (TGF-β1), decorin (DCN), and periostin was studied. The level of relative MMP-2 activation was investigated using zymography. The mRNA expression of MMP-2, TIMP-2, TGF-β1, and DCN was detected using reverse transcription polymerase chain reaction (RT-PCR), while the presence of protein was detected using immunohistochemical (IHC) and Western blot techniques. The level of MMP-2 activation was significantly elevated in tissues with Dupuytren’s contracture. RT-PCR demonstrated significantly higher expression of MMP-2, TIMP-2, TGF-β1, and DCN mRNA in the pathological tissues; and the IHC and immunoblotting studies revealed elevated expression of TGF-β1, DCN, and periostin. The balance between MMP-2 and TIMP-2 was disrupted in patients with Dupuytren’s disease. TGF-β1, DCN, and periostin are involved in extracellular matrix (ECM) homeostasis in Dupuytren’s contracture.

Ventricular remodelling in prediabetes: the heart of the matter!

proteolytic enzymes that degrade collagens, and their tissue inhibitors (TIMPS), may lead to increased degradation, contributing to ventricular dilatation and remodelling. Moreover, the role of hyperglycaemia in ventricular remodelling is emerging, and may prove to be an important pathological target, linking activation of the type β transforming growth factor (TGFβ) signalling pathway with extracellular matrix protein deposition and myocyte hypertrophy. In this issue of Experimental Physiology, D’Souza et al. (2011) present compelling evidence that the remodelling of the ventricular myocardium, which ultimately results in decompensated heart failure, occurs during mild hyperglycaemia, prior to the onset of type II diabetes. The authors modelled the prediabetic state utilizing the non-obese Goto‐Kakizaki rat, which exhibits impaired glucose tolerance and mildly elevated fasting blood glucose levels at 8 weeks of age, compared with agematched control animals. The presence of ventricular remodelling was confirmed morphologically, by myocyte hypertrophy, increased extracellular matrix deposition andfibrosis,measuredasincreasedcollagen density. Similar findings have also been identified in myocardial infarction-induced heart failure in diabetic Goto‐Kakizaki rats (Vahtola et al. 2010). D’Souza and colleagues also note a potentially interesting difference in intercalated disk configuration between Goto‐Kakizaki rats in prediabetes and control animals, which reflects altered structural contact between myocytes, but may also have a bearing on electrical coupling and conduction within the ventricle, and may warrant further attention. This study by D’Souza et al. (2011),clearlydemonstratesthatthelevelof hyperglycaemia in prediabetes is sufficient to stimulate recapitulation of the fetal gene phenotype, indicated by increased levels of brain natriuretic peptide and cellular αskeletal actin, both of which are typically upregulated in hypertensive hypertrophy and diabetes-induced hypertrophy and are associated with elevated TGFβ1 expression. Upregulation of MMP2 and MMP9 was also observed compared with control rats, consistent with the perturbation of the extracellular matrix. The MMP/TIMP family is diverse, and its expression is suggested to be region specific. Matrix metalloproteinases 2 and 9 have been shown previously to be upregulated in different facets of cardiac dysfunction, including dilated cardiomyopathy and myocardial infarction, which is supported by overexpression studies (Spinale, 2007). However, studies of ventricular pressure and volume overload suggest a more complex MMP/TIMP profile. Further work will be required to establish a fully comprehensive profile of the MMP/TIMP expression levels that exist in prediabetes and, most importantly, how they develop during disease progression. An important area in which study by D’Souza et al. (2011) is lacking is the presence of functional data to confirm ventricular impairment, and it is imperative that future studies combine mechanistic examination of the signalling processes underlying structural remodelling with functional studies.

TGF-β1 Induces an Age-Dependent Inflammation of Nerve Ganglia and Fibroplasia in the Prostate Gland Stroma of a Novel Transgenic Mouse

TGF-β1 is overexpressed in wound repair and in most proliferative disorders including benign prostatic hyperplasia and prostate cancer. The stromal microenvironment at these sites is reactive and typified by altered phenotype, matrix deposition, inflammatory responses, and alterations in nerve density and biology. TGF-β1 is known to modulate several stromal responses; however there are few transgenic models to study its integrated biology. To address the actions of TGF-β1 in prostate disorders, we targeted expression of an epitope tagged and constitutively active TGF-β1 via the enhanced probasin promoter to the murine prostate gland epithelium. Transgenic mice developed age-dependent lesions leading to severe, yet focal attenuation of epithelium, and a discontinuous basal lamina. These changes were associated with elevated fibroplasia and frequency of collagenous micronodules in collapsed acini, along with an induced inflammation in nerve ganglia and small vessels. Elevated recruitment of CD115+ myeloid cells but not mature macrophages was observed in nerve ganglia, also in an age-dependent manner. Similar phenotypic changes were observed using a human prostate epithelium tissue recombination xenograft model, where epithelial cells engineered to overexpress TGF-β1 induced fibrosis and altered matrix deposition concurrent with inflammation in the stromal compartment. Together, these data suggest that elevated TGF-β1 expression induces a fibroplasia stromal response associated with breach of epithelial wall structure and inflammatory involvement of nerve ganglia and vessels. The novel findings of ganglia and vessel inflammation associated with formation of collagenous micronodules in collapsed acini is important as each of these are observed in human prostate carcinoma and may play a role in disease progression.

Successful Incorporation of Tissue-Engineered Porcine Small-Intestinal Submucosa as Substitute Flexor Tendon Graft Is Mediated by Elevated TGF-β1 Expression in the Rabbit

Successful Incorporation of Tissue-Engineered Porcine Small-Intestinal Submucosa as Substitute Flexor Tendon Graft Is Mediated by Elevated TGF-β1 Expression in the Rabbit

Successful Incorporation of Tissue-Engineered Porcine Small-Intestinal Submucosa as Substitute Flexor Tendon Graft Is Mediated by Elevated TGF-β1 Expression in the Rabbit

Ideal tendon repair materials combine minimal donor-site morbidity and ready availability with excellent healing and postoperative function. Bioengineered porcine small-intestinal submucosa (SIS) was compared with tendon autografts as a potential human flexor tendon graft substitute. Rabbit zone II flexor digitorum profundus segments were excised in 40 rabbits. Randomized tendon repair consisted of either interposition reversed autograft or SIS, passed beneath the A2 and A4 pulleys. Forepaws were statically splinted for 3 weeks followed by unrestricted motion. Animals were killed at 7, 14, 28, and 56 days. Specimens were analyzed for hydroxyproline content (absorption spectroscopy) and tensile strength. Hematoxylin-eosin and Movat-stained sections of the central graft and distal repair site were semiquantitatively scored for total cellularity, inflammatory cell content, foreign-body reaction, vascularity, mature collagen content, and new collagen deposition. Transforming growth factor-beta (TGF-beta1) and TGF-beta1 receptor immunostaining was performed. At week 1, SIS hydroxyproline content was significantly reduced compared with autograft hydroxyproline content. However, week 2 SIS hydroxyproline content increased to equivalent values. Collagen deposition was evident in SIS by week 1 but negligible in autograft. More rapid total and inflammatory cell increases occurred in SIS by 4 weeks. A stronger early inflammatory reaction also occurred. More rapid SIS neovascularization occurred despite a greater foreign-body reaction. Small-intestinal submucosa vascularity was markedly greater at weeks 1 and 2 and equivalent thereafter. At week 4, SIS intrinsic tensile strength (suture removed) exceeded that of both autograft and suture material. Preoperative TGF-beta1 immunostaining in SIS was less than that of autograft but greater during weeks 2 and 4. Earlier neovascularization, increased TGF-beta1 levels, and increased collagen deposition, along with greater intrinsic repair strength relative to both autograft and suture strength at week 4, make SIS a promising flexor tendon graft substitute. Future studies examining tendon excursion are planned.

Fluvastatin decreases cardiac fibrosis possibly through regulation of TGF-β 1/Smad 7 expression in the spontaneously hypertensive rats

Statins ameliorate myocardial fibrosis after myocardial infarction. We designed this study to determine whether fluvastatin reduced hypertension-induced myocardial hypertrophy and fibrosis and whether these fluvastatin effects involved transforming growth factor β1 (TGF-β1) and Smad 7, factors known to play a role in the myocardial hypertrophy and fibrosis. We randomized 14 week old spontaneously hypertensive rats (SHRs) to receiving vehicle or 5–20 mg/kg/day fluvastatin for 8 weeks. Wistar Kyoto (WKY) rats receiving vehicle or 10 mg/kg/day fluvastatin were also studied. SHRs had an increased blood pressure, left ventricular hypertrophy and fibrosis compared with WKY rats. SHRs also had an elevated TGF-β1 expression and a decreased Smad 7 expression. These changes in SHRs were dose-dependently attenuated by fluvastatin. For example, the hydroxyproline content was 3.2 ± 0.1, 4.0 ± 0.1 and 3.5 ± 0.1 µg/mg heart and the Smad 7 protein expression was 5.1 ± 0.6, 1.0 ± 0.1 and 4.1 ± 0.7 arbitrary units for WKY rats, SHRs and SHRs receiving 20 mg/kg/day fluvastatin, respectively. The hydroxyproline content in the SHRs treated with or without fluvastatin was positively correlated with the left ventricular mass index, systolic blood pressure and the amount of TGF-β1 proteins and negatively correlated with the Smad 7 expression level. The left ventricular mass index was positively correlated with the systolic blood pressure. Fluvastatin did not alter the blood pressure, left ventricular mass index and collagen content of WKY rats. These results suggest that fluvastatin reduces hypertension-induced myocardial hypertrophy and fibrosis. These effects may involve an increased expression of Smad 7 and a decreased expression of TFG-β1. Our results call for clinical studies to evaluate these fluvastatin effects in hypertensive patients.

Enhanced expression of transforming growth factor-β1 in inflammatory cells, α-smooth muscle actin in stellate cells, and collagen accumulation in experimental granulomatous hepatitis caused by Toxocara canis in mice

Although toxocaral granulomatous hepatitis (TGH) characterized with a dominant-Th2 type immune response is a self-limiting disease, little is known concerning the role of fibrosis-related cytokine transforming growth factor-beta 1 (TGF-β1) in pathogenesis of TGH. A detailed histological and quantitatively immunohistochemical analysis of TGF-β1, α-smooth muscle actins (α-SMA), and collagen was performed on the liver tissues from mice infected with Toxocara canis as assessed between day 1 and 42 weeks post-infection (DPI or WPI). TGF-β1 was detected mainly in infiltrating leukocytes in lesions with strong expressions from 4 to 16 WPI. Larvae per se also exhibited strong TGF-β1-like molecule expressions in the trial. Alpha-SMA was detected predominantly in hepatic stellate cells (HSC) which surrounded the lesions with moderate expressions largely throughout the period of the entire experiment. Collagen was observed to accumulate in inflammatory lesions and biliary basement with moderate to strong expressions from 1 WPI onwards in the trial. Since many evidences have indicated that leukocytes have the potential to influence HSC by producing TGF-β1 which can affect HSC to increase collagen synthesis in various liver diseases, we may propose that persistently elevated TGF-β1 expression in infiltrating leukocytes and active HSC with marked α-SMA expressions may contribute to healing of injured sites through up-stimulation of collagen deposition; in contrast, abnormally persistent collagen accumulation may cause irreversible fibrotic injury in the TGH.

T-2 Toxin and Apoptosis

T-2 toxin is a cytotoxic fungal secondary metabolite produced by various species of Fusarium. This paper reviews the reported data about the development and/or mechanisms of T-2 toxin-induced apoptosis in the hematopoietic, lymphoid and gastrointestinal tissues, dorsal skin and fetal tissues of mice and rats. Apoptosis occurs earlier in hematopoietic tissues than in lymphoid tissues, and moreover there is a difference among lymphocyte populations in the susceptibility to T-2 toxin. C-fos plays an important role in the early phase of T-2 toxin-induced apoptosis in hematopoietic and lymphoid tissues through mobilization of [Ca2+]i and the PKC-dependent pathway, but not through Fas/Fasligand or p53-related pathways. In the gastrointestinal tract, apoptosis develops earlier in the small intestine than in the stomach, and is more prominent in the small intestine than in the large intestine. In the dorsal skin topically treated with T-2 toxin, epidermal basal cell proliferating activity is first suppressed due to the elevated TGF-β1 expression, and then basal cell apoptosis develops concomitantly with the elevation of c-fos, c-jun and TNF- α mRNAs expression. In the fetuses, T-2 toxin-induced apoptosis is mainly observed in the central nervous and skeletal systems. In the fetal brain, T-2 toxin induces oxidative stress, followed by activation of the MAPK-JNK-c-Jun pathway, finally resulting in apoptosis. The TNF receptor pathway may also be involved in T-2 toxin-induced apoptosis in the fetal brain. Thus, the development and mechanisms of T-2 toxin-induced apoptosis differ somewhat depending on the tissues affected.

Conditional tetracycline-regulated expression of TGF-β1 in liver of transgenic mice leads to reversible intermediary fibrosis

Based on the tetracycline-regulated gene expression system, a double-transgenic mouse model for liver fibrosis was established in which the expression of transforming growth factor β1 (TGF-β1) can be regulated deliberately by addition or removal of doxycycline hydrochloride to the drinking water. TGF-β1 plasma levels in induced double-transgenic mice reached values ranging from 250 to 1,200 ng/mL, being 10 to 30 times above the normal plasma levels. By applying a cyclic induction-deinduction protocol, deleterious effects of the high plasma TGF-β1 levels were overcome. By using this protocol, liver fibrosis occurred within a few cycles and progressed further to an intermediary fibrosis when cyclic induction was continued. On histochemical staining, a marked perisinusoidal deposition of extracellular matrix was detected accompanied by the activation of hepatic stellate cells as shown by alpha-smooth muscle actin (α-SMA) expression. Apoptosis of hepatocytes was prominent in TGF-β1 high producers, leading to a decreasing number of TGF-β1-expressing cells with time. No compensatory proliferation of hepatocytes could be detected. In advanced stages, fibrogenesis could be stopped by switching off TGF-β1 production and reversal of fibrosis could be shown by (immuno)histochemistry within 6 to 21 days. Determination of messenger RNA (mRNA) levels of procollagen I and III, laminin (B1), matrix metalloproteinase (MMP)-2, -9, and -13, and tissue inhibitor of matrix metalloproteinase (TIMP)-1 and -2 by real-time reverse-transcription polymerase chain reaction (RT-PCR) provided insight into some mechanistic details of the fibrogenic process and its reversal. In conclusion, this model will enable the analysis of fibrogenesis at progressive stages and help in elucidating the cellular changes during development and regression of liver fibrosis caused by elevated TGF-β1 expression. (H epatology 2003;37:1067-1078.)

Bidirectional Effect of Transforming Growth Factor-β1 (TGF-β1) and Stimulatory Effect of Platelet-Derived Growth Factor (PDGF) on Mitogenesis of Human Myometrial and Leiomyoma Cells

Objectives: TGF-β1 is the prototype of a bidirectional regulator of cell growth that can either inhibit or stimulate the proliferation of smooth muscle cells. These effects of TGF-β1 are dependent on the type and differentiation state of the target cell, the concentration of TGF-β1, and the presence of other growth factors. At least a part of TGF-β1-mediated stimulation of growth is associated with increased production of PDGF, which is another potent mitogen for smooth muscle cells. We have previously shown the elevated expression of TGF-β1 in the leiomyomatous myometrium and the increased active to latent TGF-β1 ratio under the influence of ovarian hormones. In the present study, we investigated the mitogenic effects of TGF-β1, anti-TGF-β antibody, and PDGF on myometrial and leiomyoma smooth muscle cells.