Small extracellular vesicles from irradiated lung epithelial cells promote the activation of fibroblasts in pulmonary fibrosis.

BACKGROUND: Alveolar epithelial injury and dysfunction are the risk factors for radiation-induced pulmonary fibrosis (RIPF). However, it is not clear about the relationship between RIPF and the small extracellular vesicles (sEV) secreted by irradiated alveolar epithelial cells. Based on the activation of fibroblasts, this study explored the role of sEV derived from alveolar epithelial cells in RIPF and the potential mechanisms. METHODS: Transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and western blotting were used to characterize sEV. Western blotting was used to detect fibrosis-associated proteins. Cell counts and transwell assays were used to evaluate the proliferation and migration ability of fibroblasts. RT-PCR was used to observe the extracellular matrix (ECM) synthesized by fibroblasts, miRNA changes in the sEV were determined by second-generation sequencing. RESULTS: TEM, NTA, and western blotting showed the extracellular vesicles with a double-layer membrane structure of approximately 100 nm in diameter. The sEV derived from irradiated A549, HBEC3-KT, and MLE12 cells upregulated FN1 and alpha-SMA proteins expression in fibroblasts and drove the fibroblast to myofibroblast transition, and the sEV from irradiated mouse bronchoalveolar lavage fluid (BALF) affirmed the same results. In addition, the sEV derived from irradiated alveolar epithelial cells significantly increased the migration ability of fibroblasts and the expression of extracellular matrix proteins such as FN1. The results of miRNA sequencing of sEV in BALF of rats with RIPF showed that the metabolic pathway may be important for miRNA to regulate the activation of fibroblasts. CONCLUSION: The sEV derived from radiated pulmonary epithelial cells promote the activation, migration and extracellular matrix proteins expression of lung fibroblasts; miRNA in sEV may be an important molecular that affects the activation of lung fibroblasts.

METTL1-m7 G-EGFR/EFEMP1 axis promotes the bladder cancer development.

BACKGROUND: The posttranscriptional modifications of transfer RNA (tRNA) are critical for all aspects of the tRNA function and have been implicated in the tumourigenesis and progression of many human cancers. By contrast, the biological functions of methyltransferase-like 1 (METTL1)-regulated m7 G tRNA modification in bladder cancer (BC) remain obscure. RESULTS: In this research, we show that METTL1 was highly expressed in BC, and its level was correlated with poor patient prognosis. Silencing METTL1 suppresses the proliferation, migration and invasion of BC cells in vitro and in vivo. Multi-omics analysis reveals that METTL1-mediated m7 G tRNA modification altered expression of certain target genes, including EGFR/EFEMP1. Mechanistically, METTL1 regulates the translation of EGFR/EFEMP1 via modifying certain tRNAs. Furthermore, forced expression of EGFR/EFEMP1 partially rescues the effect of METTL1 deletion on BC cells. CONCLUSIONS: Our findings demonstrate the oncogenic role of METTL1 and the pathological significance of the METTL1-m7 G-EGFR/EFEMP1 axis in the BC development, thus providing potential therapeutic targets for the BC treatment.

Re-evaluation casts doubt on the pathogenicity of homozygous USH2A p.C759F.

Mutations in USH2A are a common cause of Retinitis Pigmentosa (RP). Among the most frequently reported USH2A variants, c.2276G>T (p.C759F) has been found in both affected and healthy individuals. The pathogenicity of this variant remains controversial since it was detected in homozygosity in two healthy siblings of a Spanish family (S23), eleven years ago. The fact that these individuals remain asymptomatic today, prompted us to study the presence of other pathogenic variants in this family using targeted resequencing of 26 retinal genes in one of the affected individuals. This approach allowed us to identify one novel pathogenic homozygous mutation in exon 13 of PDE6B (c.1678C>T; p.R560C). This variant cosegregated with the disease and was absent in 200 control individuals. Remarkably, the identified variant in PDE6B corresponds to the mutation responsible of the retinal degeneration in the naturally occurring rd10 mutant mice. To our knowledge, this is the first report of the identification of the rd10 mice mutation in a RP family. These findings, together with a review of the literature, support the hypothesis that homozygous p.C759F mutations are not pathogenic and led us to exclude the implication of p.C759F in the RP of family S23. Our results indicate the need of re-evaluating all families genetically diagnosed with this mutation.

Proline/arginine-rich end leucine-rich repeat protein N-terminus is a novel osteoclast antagonist that counteracts bone loss.

(hbd) PRELP is a peptide corresponding to the N-terminal heparin binding domain of the matrix protein proline/arginine-rich end leucine-rich repeat protein (PRELP). (hbd) PRELP inhibits osteoclastogenesis entering pre-fusion osteoclasts through a chondroitin sulfate- and annexin 2-dependent mechanism and reducing the nuclear factor-κB transcription factor activity. In this work, we hypothesized that (hbd) PRELP could have a pharmacological relevance, counteracting bone loss in a variety of in vivo models of bone diseases induced by exacerbated osteoclast activity. In healthy mice, we demonstrated that the peptide targeted the bone and increased trabecular bone mass over basal level. In mice treated with retinoic acid to induce an acute increase of osteoclast formation, the peptide consistently antagonized osteoclastogenesis and prevented the increase of the serum levels of the osteoclast-specific marker tartrate-resistant acid phosphatase. In ovariectomized mice, in which osteoclast activity was chronically enhanced by estrogen deficiency, (hbd) PRELP counteracted exacerbated osteoclast activity and bone loss. In mice carrying osteolytic bone metastases, in which osteoclastogenesis and bone resorption were enhanced by tumor cell-derived factors, (hbd) PRELP reduced the incidence of osteolytic lesions, both preventively and curatively, with mechanisms involving impaired tumor cell homing to bone and tumor growth in the bone microenvironment. Interestingly, in tumor-bearing mice, (hbd) PRELP also inhibited breast tumor growth in orthotopic sites and development of metastatic disease in visceral organs, reducing cachexia and improving survival especially when administered preventively. (hbd) PRELP was retained in the tumor tissue and appeared to affect tumor growth by interacting with the microenvironment rather than by directly affecting the tumor cells. Because safety studies and high-dose treatments revealed no adverse effects, (hbd) PRELP could be employed as a novel biological agent to combat experimentally induced bone loss and breast cancer metastases, with a potential translational impact.

Reduction in facial photodamage by a topical growth factor product.

A topical gel containing a proprietary mixture of over 110 growth factors, cytokines, and soluble matrix proteins secreted by human dermal fibroblasts was evaluated for safety and efficacy in the treatment of mild to severe facial photodamage. In a double-blind study, 60 subjects were randomly assigned to receive either active gel or the vehicle and applied twice daily for 6 months along with a moisturizing cleanser and sunscreen. Efficacy (profilometry, photography, and clinical assessment) and safety (adverse event reporting) measures were evaluated at 0, 3, and 6 months. Treatment with the active gel for 3 months produced greater reduction in fine lines and wrinkles than the vehicle treatment as measured by objective and subjective assessment techniques. The results were either statistically significant (P < or = .05) or trending towards statistical significance (P < or = .1). This study demonstrates that addition of a topical formulation of growth factors and cytokines to a basic skin care regimen reduces the signs of photoaging.

Cartilage oligomeric matrix protein induction of chronic arthritis in mice.

OBJECTIVE: To develop a new mouse model for arthritis using cartilage oligomeric matrix protein (COMP) and to study the role of major histocompatibility complex (MHC) and Ncf1 genes in COMP-induced arthritis (COMPIA). METHODS: Native (pentameric) and denatured (monomeric) COMP purified from a rat chondrosarcoma was injected into mice with Freund's adjuvant to induce arthritis. C3H.NB, C3H.Q, B10.P, B10.Q, (B10.Q x DBA/1)F1, (BALB/c x B10.Q)F1, Ncf1 mutated, H-2Aq, H-2Ap, and human DR4+-transgenic mice were used. Anti-COMP antibodies and COMP levels in the immune sera were analyzed, and passive transfer of arthritis with purified immune sera was tested. RESULTS: Immunization with rat COMP induced a severe, chronic, relapsing arthritis, with a female preponderance, in the mice. The disease developed in C3H.NB mice, but not in B10.P mice, although they share the same MHC haplotype. Both H-2q and H-2p MHC haplotypes allowed the initiation of COMPIA. Using H-2Aq-transgenic and H-2Ap-transgenic mice, we demonstrated a role of both the Aq and Ep class II molecules in this model. Interestingly, the introduction of a mutation in the Ncf1 gene, which is responsible for the reduced oxidative burst phenotype, into the COMPIA-resistant B10.Q mouse strain rendered them highly susceptible to arthritis. In addition, the transfer of anti-COMP serum was found to induce arthritis in naive mice. Mice transgenic for the rheumatoid arthritis (RA)-associated DR4 molecule were found to be highly susceptible to COMPIA. CONCLUSION: Using rat COMP, we have developed a new and unique mouse model of chronic arthritis that resembles RA. This model will be useful as an appropriate and alternative model for studying the pathogenesis of RA.

RHAMM-R3 peptide vaccination in patients with acute myeloid leukemia, myelodysplastic syndrome, and multiple myeloma elicits immunologic and clinical responses.

The receptor for hyaluronic acid-mediated motility (RHAMM) is an antigen eliciting both humoral and cellular immune responses in patients with acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), and multiple myeloma (MM). We initiated a phase 1 clinical trial vaccinating 10 patients with R3 (ILSLELMKL), a highly immunogenic CD8(+) T-cell epitope peptide derived from RHAMM. In 7 of 10 patients, we detected an increase of CD8(+)/HLA-A2/RHAMM R3 tetramer(+)/CD45RA(+)/CCR7(-)/CD27(-)/CD28(-) effector T cells in accordance with an increase of R3-specific CD8(+) T cells in enzyme linked immunospot (ELISpot) assays. In chromium release assays, a specific lysis of RHAMM-positive leukemic blasts was shown. Three of 6 patients with myeloid disorders (1/3 AML, 2/3 MDS) achieved clinical responses: one patient with AML and one with MDS showed a significant reduction of blasts in the bone marrow after the last vaccination. One patient with MDS no longer needed erythrocyte transfusions after 4 vaccinations. Two of 4 patients with MM showed a reduction of free light chain serum levels. Taken together, RHAMM-R3 peptide vaccination induced both immunologic and clinical responses, and therefore RHAMM constitutes a promising target for further immunotherapeutic approaches. This study is registered at http://ISRCTN.org as ISRCTN32763606 and is registered with EudraCT as 2005-001706-37.

Effect of amelogenin extracellular matrix protein and compression on hard-to-heal venous leg ulcers.

OBJECTIVE: To compare hard-to-heal venous leg ulcers treated with compression therapy alone versus compression therapy with amelogenin protein. Parameters used were: percentage reduction in wound size, number of improved ulcers, pain related to the disease and at dressing changes, amount and nature of exudate, and the safety and tolerability of the two treatments. METHOD: This was an open randomised comparative parallel group multicentre investigation with a three-week run-in period. Inclusion criteria included adult, mobile patients with hard-to-heal venous leg ulcers that had been treated with compression therapy for at least one month prior to screening. The ulcers had to be at least six months old, with a surface area at inclusion of 10-30cm2, and not demonstrating excessive exudate or signs of infection. At the end of the run-in period, additional criteria for eligibility, such as change in wound area of +/- > or = 50% and a wound area between 8cm2 and 36cm2 were applied. Patients were randomised to treatment with amelogenin plus high compression bandaging or high compression bandaging alone. All participants received high compression bandaging therapy one month prior to and during the three-week run-in period, as well as throughout the 12 weeks of active treatment. RESULTS: Eighty-three patients were randomised and received treatment: 42 with high compression plus amelogenin (amelogenin group) and 41 to high compression therapy alone (control group). The amelogenin group had a greater percentage reduction in ulcer size (mean - 33.1%) compared with the control group (mean - 11.07%) from baseline to the last visit (p = 0.06). The number of improved ulcers was significantly greater (p = 0.01) in the amelogenin group than in the control group. Compensating for baseline characteristics by multiple regression resulted in a statistically significant (p = 0.03) larger reduction in change in ulcer size in the amelogenin group. Statistically significant differences in favour of the amelogenin group were also found for reduction in ulcer-related pain (p = 0.01), reduction in pain at dressing changes (p = 0.02) and the proportion of patients with 'none' or 'low' levels of exudate (p = 0.01). CONCLUSION: The combination of amelogenin with high compression promotes the healing process in hard-to-heal ulcers. Application of amelogenin as an adjunct to compression results in a significant reduction in ulcer size, improvement in the state of ulcers, reduced pain and a larger proportion of ulcers with low levels of exudate. The results of this study are statistically and clinically significant.