Long-term efficacy and safety of tildrakizumab for moderate-to-severe psoriasis: pooled analyses of two randomized phase III clinical trials (reSURFACE 1 and reSURFACE 2) through 148 weeks.

BACKGROUND: Tildrakizumab is a specific anti-interleukin-23p19 monoclonal antibody approved for the treatment of plaque psoriasis. OBJECTIVES: To evaluate the long-term efficacy and safety of tildrakizumab treatment for patients with moderate-to-severe psoriasis for up to 148 weeks. METHODS: Pooled analysis from two double-blind, randomized controlled trials: reSURFACE 1 and reSURFACE 2. Efficacy was assessed for responders (≥ 75% improvement in Psoriasis Area and Severity Index; PASI 75) and partial responders (PASI 50-75) to tildrakizumab 100 mg and 200 mg at week 28 who were maintained on the same dose (administered every 12 weeks), and for partial responders or nonresponders (PASI < 50) to etanercept 50 mg at week 28 who, after a 4-week washout, were switched to tildrakizumab 200 mg (administered at weeks 32 and 36, and every 12 weeks thereafter). Safety was assessed in the all-patients-as-treated population. Three different methods of imputing missing data were used: nonresponder imputation (NRI), multiple imputation and observed cases. The Clinicaltrials.gov numbers are NCT01722331 (reSURFACE 1) and NCT01729754 (reSURFACE 2). RESULTS: At week 148 (NRI), 72·6%, 53·8% and 28·9% of tildrakizumab 100-mg responders and 80·2%, 59·9% and 32·6% of tildrakizumab 200-mg responders had PASI 75, 90 and 100 responses, respectively. For partial responders to tildrakizumab 100 mg and 200 mg, the proportions of patients achieving PASI 75, 90 and 100 responses were 32·5%, 25·0% and 10·0%; and 47·1%, 27·5% and 12·8%, respectively. For patients who were partial responders or nonresponders to etanercept, the proportions of patients achieving PASI 75, 90 and 100 responses were 66·9%, 43·8% and 14·9% at week 148. Rates of discontinuations due to adverse events [tildrakizumab 100 mg: 1·7 per 100 patient-years (PYs); tildrakizumab 200 mg: 1·2 per 100 PYs] and exposure-adjusted rates of serious adverse events (5·9 per 100 PYs; 5·5 per 100 PYs), severe infections (1·1 per 100 PYs; 1·1 per 100 PYs), malignancies (0·6 per 100 PYs; 0·4 per 100 PYs) and major adverse cardiovascular events (0·4 per 100 PYs; 0·5 per 100 PYs) were low. CONCLUSIONS: Tildrakizumab was well tolerated and efficacy was well maintained in week 28 responders who continued tildrakizumab treatment through 3 years, or improved among etanercept partial responders or nonresponders who switched to tildrakizumab. What's already known about this topic? Tildrakizumab 100 mg and 200 mg are efficacious and well tolerated with short-term use in the treatment of patients with moderate-to-severe plaque psoriasis. What does this study add? High levels of efficacy are maintained for up to 3 years of psoriasis treatment with tildrakizumab. There is a favourable long-term safety profile with both tildrakizumab 100 mg and 200 mg, with a low incidence of adverse events of special interest through 3 years.

Diffuse Pulmonary Meningotheliomatosis with Sarcomatous Transformation in a Shiba Dog.

A 2-year-old neutered female Shiba dog exhibited laboured breathing for 1 month. Computed tomography of the thoracic cavity revealed multiple nodules (2-5 mm diameter) in the lungs. Grossly, the lungs were firm and normal in shape. The nodules were grey-white in colour. Microscopically, the nodules were non-encapsulated and exhibited an irregular shape. They were composed of polygonal or spindle cells with indistinct cell borders arranged in sheets. The cells had large, round, hyperchromatic nuclei and abundant pale eosinophilic cytoplasm with no atypia. Intrapulmonary arterial emboli and infiltration into the bronchioles were observed. Immunohistochemically, the cells were positive for vimentin and negative for cytokeratin, glial fibrillary acidic protein and α-smooth muscle actin. Ultrastructurally, the cells displayed cytoplasmic processes, desmosomes and intermediate filaments. These findings led to a diagnosis of diffuse pulmonary meningotheliomatosis with sarcomatous transformation. To the best of our knowledge, this is the first report of diffuse pulmonary meningotheliomatosis in a dog.

Prognostic significance of hypermethylation of death-associated protein kinase (DAPK) gene CpG island in dogs with high-grade B-cell lymphoma.

Death-associated protein kinase (DAPK) is a serine/threonine kinase and a tumour suppressor gene. Diffuse large B-cell lymphomas with inactivated DAPK through hypermethylation of a CpG island is known to result in a biologically aggressive phenotype in humans. This retrospective study was carried out to analyse the prognostic significance of DAPK CpG island hypermethylation in canine lymphoma. We hypothesized that DAPK CpG island hypermethylation can be a negative prognostic indicator in dogs with nodal high-grade B-cell lymphoma. Forty-seven dogs with high-grade B-cell lymphoma, according to the updated Kiel classification, were evaluated after being treated with a CHOP (vincristine, cyclophosphamide, doxorubicin and prednisolone)-based chemotherapy protocol. The methylation status of the DAPK CpG island was examined by methylation-specific PCR. Progression-free survival (PFS) and overall survival (OS) were compared using the Kaplan-Meier analysis and log-rank test. The cox proportional hazard regression model was used to evaluate the effect of multiple variables. Hypermethylation of the DAPK CpG island was detected in 21 of the 47 dogs. The PFS and OS in dogs with the hypermethylation (median: 220 and 266 days, respectively) were significantly shorter than those of dogs without hypermethylation (median: 301 and 412 days, respectively) (PFS, P = .036; OS, P = .007). In the multivariate analysis, hypermethylation of the DAPK CpG island remained an independent prognostic factor in predicting shortened PFS (P = .047) and OS (P = .021) as well as clinical substage b. Overall, hypermethylation of the DAPK CpG island was a negative prognostic factor in canine high-grade B-cell lymphoma.

Cancer screening for older people: to screen or not to screen.

In this scoping review, the evidence of the benefits of screening older people for the five most common types of cancer in Hong Kong, namely colorectal, lung, breast, liver, and prostate cancers, is discussed. Although cancer treatments can be extensive and a good prognosis is less likely if cancer is diagnosed at a late stage, screening programmes for older people in primary care remain a matter of contention. The general recommendation for the screening of older people is to adopt an individualised approach that takes account of not only age but also co-morbidity, life expectancy, harms and benefits, and patient's preference.

Characterization of PTEN mutations in brain cancer reveals that pten mono-ubiquitination promotes protein stability and nuclear localization.

PTEN is a PIP3 phosphatase that antagonizes oncogenic PI3-kinase signalling. Due to its critical role in suppressing the potent signalling pathway, it is one of the most mutated tumour suppressors, especially in brain tumours. It is generally thought that PTEN deficiencies predominantly result from either loss of expression or enzymatic activity. By analysing PTEN in malignant glioblastoma primary cells derived from 16 of our patients, we report mutations that block localization of PTEN at the plasma membrane and nucleus without affecting lipid phosphatase activity. Cellular and biochemical analyses as well as structural modelling revealed that two mutations disrupt intramolecular interaction of PTEN and open its conformation, enhancing polyubiquitination of PTEN and decreasing protein stability. Moreover, promoting mono-ubiquitination increases protein stability and nuclear localization of mutant PTEN. Thus, our findings provide a molecular mechanism for cancer-associated PTEN defects and may lead to a brain cancer treatment that targets PTEN mono-ubiquitination.

Genetic and epigenetic aberrations of p16 in feline primary neoplastic diseases and tumor cell lines of lymphoid and non-lymphoid origins.

The p16 gene acts as a tumor suppressor by regulating the cell cycle and is frequently inactivated in human and canine cancers. The aim of this study was to characterize genetic and epigenetic alterations of the p16 in feline lymphoid and non-lymphoid malignancies, using 74 primary tumors and 11 tumor cell lines. Cloning of feline p16 and subsequent sequence analysis revealed 11 germline sequence polymorphisms in control cats. Bisulfite sequencing analysis of the p16 promoter region in a feline lymphoma cell line revealed that promoter methylation was associated with decreased mRNA expression. Treatment with a demethylating agent restored mRNA expression of the silenced p16. PCR amplification and sequencing analysis detected homozygous loss (five tumors, 6.7%) and a missense mutation (one tumor, 1.4%) in the 74 primary tumors analyzed. Methylation-specific PCR analysis revealed promoter methylation in 10 primary tumors (14%). Promoter methylation was frequent in B cell lymphoid tumors (7/21 tumors, 33%). These genetic and epigenetic alterations were also observed in lymphoma and mammary gland carcinoma cell lines, but not detected in non-neoplastic control specimens. These data indicate that molecular alterations of the p16 locus may be involved in the development of specific types of feline cancer, and warrant further studies to evaluate the clinical value of this evolutionarily-conserved molecular alteration in feline cancers.

Changes in Foxp3-Positive Regulatory T Cell Number in the Intestine of Dogs With Idiopathic Inflammatory Bowel Disease and Intestinal Lymphoma.

Although regulatory T cells (Tregs) play an integral role in immunologic tolerance and the maintenance of intestinal homeostasis, their involvement in canine gastrointestinal diseases, including idiopathic inflammatory bowel disease (IBD) and intestinal lymphoma, remains unclear. Here we show altered numbers of forkhead box P3 (Foxp3)-positive Tregs in the intestine of dogs with IBD and intestinal lymphoma. IBD was diagnosed in 48 dogs; small cell intestinal lymphoma was diagnosed in 46 dogs; large cell intestinal lymphoma was diagnosed in 30 dogs; and 25 healthy beagles were used as normal controls. Foxp3-positive Tregs in the duodenal mucosa were examined by immunohistochemistry and immunofluorescence. Duodenal expression of interleukin-10 mRNA was quantified by real-time reverse transcription polymerase chain reaction. The number of Foxp3-positive lamina propria cells and the expression of interleukin-10 mRNA were significantly lower in dogs with IBD than in healthy dogs and dogs with intestinal lymphoma. The number of Foxp3-positive intraepithelial cells was higher in dogs with small cell intestinal lymphoma. Some large cell intestinal lymphoma cases had high numbers of Foxp3-positive cells, but the increase was not statistically significant. Double-labeling immunofluorescence showed that CD3-positive granzyme B-negative helper T cells expressed Foxp3. In small cell intestinal lymphoma cases, the overall survival of dogs with a high Treg density was significantly worse than that of dogs with a normal Treg density. These results suggest that a change in the number of Foxp3-positive Tregs contributes to the pathogenesis of canine IBD and intestinal lymphoma by disrupting mucosal tolerance and suppressing antitumor immunity, respectively.

Clinical evaluation of the additive effect of diquafosol tetrasodium on sodium hyaluronate monotherapy in patients with dry eye syndrome: a prospective, randomized, multicenter study.

PURPOSE: To assess the additive effect of diquafosol tetrasodium on sodium hyaluronate monotherapy in patients with dry eye syndrome. METHODS: This study evaluated 64 eyes of 32 patients (age: 62.6±12.8 years (mean±SD)) in whom treatment with 0.1% sodium hyaluronate was insufficiently responsive. The eyes were randomly assigned to one of the two regimens in each patient: topical administration of sodium hyaluronate and diquafosol tetrasodium in one eye, and that of sodium hyaluronate in the other. Before treatment, and 2 and 4 weeks after treatment, we determined tear volume, tear film break-up time (BUT), fluorescein and rose bengal vital staining scores, subjective symptoms, and adverse events. RESULTS: We found a significant improvement in BUT (P=0.049, Dunnett test), fluorescein and rose bengal staining scores (P=0.02), and in subjective symptoms (P=0.004 for dry eye sensation, P=0.02 for pain, and P=0.02 for foreign body sensation) 4 weeks after treatment in the diquafosol eyes. On the other hand, we found no significant change in these parameters after treatment in the control eyes. CONCLUSIONS: In dry eyes, where sodium hyaluronate monotherapy was insufficient, diquafosol tetrasodium was effective in improving objective and subjective symptoms, suggesting its viability as an option for the additive treatment of such eyes.

Anti-host isohemagglutinin production is associated with a higher risk of acute GVHD in ABO-incompatible transplantation.

We retrospectively analyzed the association between anti-host isohemagglutinin (IH) production and the development of acute GVHD. Of 189 patients who received minor or major/minor ABO-incompatible hematopoietic SCT (HSCT) at our hospital, 36 patients (19%) showed IH production. IH was detected before the onset of acute GVHD in 10, around the same time in 8, and after the onset of acute GVHD in 17 patients. The cumulative incidence of grade II-IV acute GVHD was significantly higher in the IH+ group compared with the IH- group (P<0.0001). The higher risk of acute GVHD that was associated with IH production occurred irrespective of human leukocyte Ag compatibility and donor type. Furthermore, the incidence of acute GVHD in the IH- group was comparable to that seen in major ABO-incompatible or -compatible HSCT. Our findings not only showed a strong association between IH production and acute GVHD development, but also suggested that IH production might be a useful predictor of subsequent acute GVHD after ABO-incompatible HSCT.

Evaluation of pupil diameter after posterior chamber phakic intraocular lens implantation.

PURPOSE: To observe the changes of pupil size over time after Visian Implantable Collamer Lens (ICL) implantation. PATIENTS AND METHODS: We retrospectively examined 30 eyes of 23 consecutive patients undergoing ICL implantation. We measured the entrance and real pupil diameters using a Hartmann-Shack aberrometer (KR-9000, Topcon, Tokyo, Japan) before and 1 day, 1 week, and 1, 3, 6, and 12 months after surgery. We also investigated its relationship with the amount of vaulting using slit-lamp microscopy 1 year postoperatively. RESULTS: The entrance pupil diameters were 6.24+/-0.66 (mean+/-standard deviation) mm preoperatively, and 5.53+/-0.69, 6.18+/-0.61, 6.21+/-0.80, 6.29+/-0.74, 6.23+/-0.76, and 6.40+/-0.70 mm, 1 day, 1 week, and 1, 3, 6, and 12 months postoperatively, respectively; and the respective real pupil diameters were 5.44+/-0.55 mm preoperatively, and 4.95+/-0.60, 5.53+/-0.52, 5.55+/-0.69, 5.63+/-0.64, 5.57+/-0.64, and 5.72+/-0.60 mm, 1 day, 1 week, and 1, 3, 6, and 12 months postoperatively. Pupil diameters and the amount of vaulting were not significantly associated (Pearson correlation coefficient r=0.14, P=0.45 for entrance pupil, r=0.13, P=0.49 for real pupil). CONCLUSIONS: Both pupil diameters decreased transiently 1 day after ICL implantation, but soon recovered to the preoperative level, indicating that intraoperative mechanical irritation of the uveal tissue and the early postoperative inflammatory response may induce transient decreases in pupil diameter, and that this surgical technique alone with appropriate ICL size selection probably induces no significant pupil diameter change.

Clinical outcomes and patient satisfaction after Visian Implantable Collamer Lens removal and phacoemulsification with intraocular lens implantation in eyes with induced cataract.

PURPOSE: To assess the clinical outcomes and patient satisfaction after simultaneous Implantable Collamer Lens (ICL) removal and phacoemulsification with intraocular lens (IOL) implantation in ICL-implanted eyes with developing cataracts. METHODS: We retrospectively reviewed the clinical charts of 10 eyes of eight patients who developed significant cataract after ICL implantation. Patient age was 47.2+/-5.9 (mean+/-SD) years (range, 37-57 years). We determined visual acuity (log MAR), manifest refraction, endothelial cell density, and subjective satisfaction with visual outcomes measured using a visual analogue scale, that ranged from 0 (very dissatisfied) to 10 (very satisfied), before and 3 months after cataract surgery. RESULTS: Log MAR best spectacle-corrected visual acuity was significantly improved from 0.19+/-0.30 preoperatively to -0.06+/-0.07 postoperatively (P=0.007, Wilcoxon signed-rank test). Eight (80%) and nine (90%) of the 10 eyes were within 0.5 D (diopter) and 1.0 D of the targeted correction, respectively. The endothelial cell density was 2584.9+/-266.2 cells/mm(2) preoperatively, and 2340.1+/-269.7 cells/mm(2) postoperatively. The overall satisfaction with visual outcomes was scored 1.8+/-1.1 (range: 0-3) preoperatively, and 7.9+/-1.4 (range: 6-10) postoperatively. No vision-threatening complications were seen throughout the follow-up period. CONCLUSIONS: Simultaneous ICL removal and phacoemulsification with IOL implantation was safe and effective with predictable refractive results, and thus yielded a high level of patient satisfaction with ICL-induced cataract treatment.

Surgically induced astigmatism after posterior chamber phakic intraocular lens implantation.

AIM: To assess astigmatism induced after phakic intraocular lens (Visian ICL, STAAR Surgical) implantation. METHODS: Seventy-three eyes of 47 patients undergoing ICL implantation through a horizontal 3.0 mm clear corneal incision were retrospectively examined. The amount of corneal astigmatism before and 3 months after surgery using an automated keratometer (ARK-700A, Nidek) and corneal topography (ATRAS995, Carl Zeiss Meditec) were quantitatively investigated. The surgically induced astigmatism was assessed by vector analysis using the Holladay-Cravy-Koch formula. RESULTS: The corneal astigmatism was significantly increased from 1.10 (0.51) dioptres (D) to 1.44 (0.57) D using the keratometer (Wilcoxon signed-rank test, p<0.001). It was also significantly increased from 1.16 (0.53) D to 1.45 (0.57) D using corneal topography (p<0.001). On the other hand, the manifest astigmatism was significantly decreased from 0.93 (0.60) D to 0.72 (0.58) D (p<0.001). The surgically induced astigmatism was 0.45 (0.26) D at an axis of 93.3 degrees using the keratometer and 0.49 (0.26) D at an axis of 98.0 degrees using corneal topography. CONCLUSIONS: ICL implantation induces corneal astigmatism through a with-the-rule astigmatic shift of approximately 0.5 D, which was small but not negligible for candidates for refractive surgery.

Ocular surface and tear functions after topical cyclosporine treatment in dry eye patients with chronic graft-versus-host disease.

We investigated the effect of 0.05% topical cyclosporine (Cys) on the ocular surface and tear functions in dry eye patients with chronic GVHD (cGVHD) in a prospective comparative study. Thirty eyes of 15 patients refractory to baseline treatment were recruited and the patients assigned for topical Cys treatment group (14 eyes of 7 patients) and control group (12 eyes of 6 patients) respectively. Two patients dropped out because of intolerable irritation while using topical Cys eye drops. Visual analog scale symptom scores, corneal sensitivity, Schirmer I test value, tear film break-up time (TBUT), tear evaporation rate and ocular surface vital staining scores were recorded at baseline and at the end of the following one month. Conjunctival impression and brush cytology were performed before and after the treatment. After topical Cys treatment, significant improvements were found in symptom scores, corneal sensitivity, tear evaporation rate, TBUT, vital staining scores, goblet cells density, conjunctival squamous metaplasia grade, inflammatory cell numbers and the MUC5AC expression. Our study suggests that 0.05% topical Cys may be an effective treatment for dry eye patients with cGVHD. The improvements in the ocular surface and tear functions resulted presumably from the decreased inflammation, increased goblet cell density and MUC5AC mRNA expression. Bone Marrow Transplantation (2008) 41, 293-302; doi:10.1038/sj.bmt.1705900; published online 5 November 2007.

The differences of tear function and ocular surface findings in patients with atopic keratoconjunctivitis and vernal keratoconjunctivitis.

BACKGROUND: The pathogenesis of the ocular surface disease in atopic keratoconjunctivitis (AKC) and vernal keratoconjunctivitis (VKC) has not been fully understood. We tried to clarify the differences in the ocular surface status in patients with AKC, VKC, and healthy control subjects. METHODS: Twenty-four eyes of 12 AKC patients, 12 eyes of six VKC patients, and 20 eyes of 10 normal control subjects were studied. The subjects underwent corneal sensitivity measurements, Schirmer test, tear film break-up time (BUT), vital staining of the ocular surface, conjunctival impression and brush cytology. Impression cytology samples underwent periodic acid Schiff staining for goblet cell density, squamous metaplasia grading, and immunohistochemical staining for MUC1, 2, 4, and 5AC. Brush cytology specimens underwent staining for inflammatory cell counting and Real Time PCR for MUC1, 2, 4, and 5AC mRNA expression. RESULTS: The mean BUT, corneal sensitivity, and conjunctival goblet cell density values in AKC patients were significantly lower compared with VKC patients and control subjects. The squamous metaplasia grades in eyes with AKC were significantly higher compared to eyes with VKC and controls. The inflammatory cell response in brush cytology specimens was different between patients with AKC and VKC. Eyes with AKC showed significantly higher MUC1, 2 and 4 and lower MUC5AC mRNA expression compared to eyes with VKC. CONCLUSIONS: Differences of the infiltrates, higher level of tear instability, lower corneal sensitivity, up-regulation of MUC1, 2, and 4, and down regulation of MUC5AC were important differential features of the ocular surface disease in AKC compared with VKC.

Identification of marker genes distinguishing human periodontal ligament cells from human mesenchymal stem cells and human gingival fibroblasts.

BACKGROUND AND OBJECTIVE: Molecular gene markers, which can distinguish human bone marrow mesenchymal stem cells from human fibroblasts, have recently been reported. Messenger RNA levels of tissue factor pathway inhibitor-2, major histocompatibility complex-DR-alpha, major histocompatibility complex-DR-beta, and neuroserpin are higher in human bone marrow mesenchymal stem cells than in human fibroblasts. However, human bone marrow mesenchymal stem cells express less apolipoprotein D mRNA than human fibroblasts. Periodontal ligament cells are a heterogeneous cell population including fibroblasts, mesenchymal stem cells, and progenitor cells of osteoblasts or cementoblasts. The use of molecular markers that distinguish human bone marrow mesenchymal stem cells from human fibroblasts may provide insight into the characteristics of human periodontal ligament cells. In this study, we compared the molecular markers of human periodontal ligament cells with those of human bone marrow mesenchymal stem cells and human gingival fibroblasts. MATERIAL AND METHODS: The mRNA expression of the molecular gene markers was analyzed using real-time polymerase chain reaction. Statistical differences were determined with the two-sided Mann-Whitney U-test. RESULTS: Messenger RNA levels of major histocompatibility complex-DR-alpha and major histocompatibility complex-DR-beta were lower and higher, respectively, in human periodontal ligament cells than in human bone marrow mesenchymal stem cells or human gingival fibroblasts. Human periodontal ligament cells showed the lowest apolipoprotein D mRNA levels among the three types of cells. CONCLUSION: Human periodontal ligament cells may be distinguished from human bone marrow mesenchymal stem cells and human gingival fibroblasts by the genes for apolipoprotein D, major histocompatibility complex-DR-alpha, and major histocompatibility complex-DR-beta.

Combined detection and genotyping of Chikungunya virus by a specific reverse transcription-polymerase chain reaction.

A reverse transcription-polymerase chain reaction (RT-PCR) was developed for the detection of Chikungunya virus infection. Based on the nonstructural protein 1 (nsP1) and glycoprotein E1 (E1) genes of Chikungunya, two primer sets were designed. Total RNA were extracted from the cell culture fluid of Aedes albopictus C6/36 cells inoculated with the S27 prototype virus, isolated in Tanzania in 1953, and the Malaysian strains (MALh0198, MALh0298, and MALh0398), isolated in Malaysia in 1998. For both sets of RNA samples, the expected 354- and 294-base pair (bp) cDNA fragments were amplified effectively from the nsP1 and E1 genes, respectively. Phylogenetic analysis was conducted for the Malaysian strain and other virus strains isolated from different regions in the world endemic for Chikungunya, using partial E1 gene sequence data. The Malaysian strains isolated during the epidemics of 1998 fell into a cluster with other members of the Asian genotype.

Connective tissue growth factor is directly related to liver fibrosis.

BACKGROUND/AIMS: Connective tissue growth factor is a mitogenic, chemotactic, and cell matrix-inducing factor for fibroblasts, and is exclusively and directly induced by transforming growth factor-beta 1. In skin fibrosis, connective tissue growth factor is thought to be a downstream effector of transforming growth factor-beta 1 that is directly involved in the proliferation of connective tissue cells and the accumulation of matrix. Our aim is to confirm the involvement of connective tissue growth factor in liver fibrosis. METHODOLOGY: Specimens obtained from autopsy of 12 patients with normal liver (n = 3), hepatocellular carcinoma (n = 3), liver cirrhosis secondary to alcoholic abuse (n = 3), or viral hepatitis (n = 3), were used for digoxigenin labeled in situ hybridization. RESULTS: Abundant connective tissue growth factor messages were detected in the fibrotic area between cirrhotic nodules. Hepatocytes did not show any signals even when they became carcinomas. Normal livers showed few or no signals. CONCLUSIONS: These results confirmed the direct relationship between connective tissue growth factor gene expression and liver fibrotic change.

Characterization of the increase in bone 66 kDa protein component with healing rat fractures: stimulatory effect of zinc.

The characterization of protein components produced from bone tissues with fracture healing was investigated. Weanling rats were sacrificed between 1 and 7 days after the femoral fracture. Protein content in the femoral-diaphyseal tissues was markedly elevated by fracture healing. Moreover, when the femoral-diaphyseal tissues with fracture healing were cultured for 24 h in a serum-free medium, many proteins in the bone tissues were released into the medium. Analysis with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that many protein molecules were released from the diaphyseal tissues with fracture healing. Especially, a protein molecule of approximately 66 kDa was markedly increased by fracture healing. This protein molecule was significantly increased, when the diaphyseal tissues with fracture healing were cultured in the presence of zinc acexamate (10(-6)-10(-4) M). Zinc acexamate (10(-4) M)-induced increase in medium 66 kDa protein molecule was significantly inhibited in the presence of actinomycin D (10(-7) M) or cycloheximide (10(-6) M). The zinc effect was completely blocked in the presence of PD98059 (10(-5) M), an inhibitor of MAPK kinase, or staurosporine (10(-6) M), an inhibitor of protein kinase C. The medium 66 kDa protein molecule was significantly elevated in the presence of parathyroid hormone (1-34) (10(-7) M), insulin-like growth factor-I (10(-8) M) or transforming growth factor-beta (10(-11) M), while 17beta-estradiol (10(-9) M) did not have an effect. The effect of these bone-stimulating factors was equal to the zinc effect. Zinc did not significantly enhance the effect of insulin-like growth factor-I in increasing medium 66 kDa protein molecule. The present study demonstrates that fracture healing increases production of the approximately 66 kDa protein molecule which is a major component produced from femoral-diaphyseal tissues of weanling rats, and that this elevation is enhanced by zinc treatment.