Loss of IGFBP2 mediates alveolar type 2 cell senescence and promotes lung fibrosis.

Accumulation of senescent cells contributes to age-related diseases including idiopathic pulmonary fibrosis (IPF). Insulin-like growth factor binding proteins (IGFBPs) regulate many biological processes; however, the functional contributions of IGFBP2 in lung fibrosis remain largely unclear. Here, we report that intranasal delivery of recombinant IGFBP2 protects aged mice from weight loss and demonstrated antifibrotic effects after bleomycin lung injury. Notably, aged human-Igfbp2 transgenic mice reveal reduced senescence and senescent-associated secretory phenotype factors in alveolar epithelial type 2 (AEC2) cells and they ameliorated bleomycin-induced lung fibrosis. Finally, we demonstrate that IGFBP2 expression is significantly suppressed in AEC2 cells isolated from fibrotic lung regions of patients with IPF and/or pulmonary hypertension compared with patients with hypersensitivity pneumonitis and/or chronic obstructive pulmonary disease. Altogether, our study provides insights into how IGFBP2 regulates AEC2-cell-specific senescence and that restoring IGFBP2 levels in fibrotic lungs can prove effective for patients with IPF.

Targeted Inhibition of DNA-PKcs, ATM, ATR, PARP, and Rad51 Modulate Response to X Rays and Protons.

Small molecule inhibitors are currently in preclinical and clinical development for the treatment of selected cancers, particularly those with existing genetic alterations in DNA repair and DNA damage response (DDR) pathways. Keen interest has also been expressed in combining such agents with other targeted antitumor strategies such as radiotherapy. Radiotherapy exerts its cytotoxic effects primarily through DNA damage-induced cell death; therefore, inhibiting DNA repair and the DDR should lead to additive and/or synergistic radiosensitizing effects. In this study we screened the response to X-ray or proton radiation in cell lines treated with DDR inhibitors (DDRis) targeting ATM, ATR, DNA-PKcs, Rad51, and PARP, with survival metrics established using clonogenic assays. We observed that DDRis generate significant radiosensitization in cancer and primary cells derived from normal tissue. Existing genetic defects in cancer cells appear to be an important consideration when determining the optimal inhibitor to use for synergistic combination with radiation. We also show that while greater radiosensitization can be achieved with protons (9.9 keV/µm) combined with DDRis, the relative biological effectiveness is unchanged or in some cases reduced. Our results indicate that while targeting the DDR can significantly radiosensitize cancer cells to such combinations, normal cells may also be equally or more severely affected, depending on the DDRi used. These data highlight the importance of identifying genetic defects as predictive biomarkers of response for combination treatment.

An empirical model of proton RBE based on the linear correlation between x-ray and proton radiosensitivity.

BACKGROUND: Proton relative biological effectiveness (RBE) is known to depend on physical factors of the proton beam, such as its linear energy transfer (LET), as well as on cell-line specific biological factors, such as their ability to repair DNA damage. However, in a clinical setting, proton RBE is still considered to have a fixed value of 1.1 despite the existence of several empirical models that can predict proton RBE based on how a cell's survival curve (linear-quadratic model [LQM]) parameters α and ß vary with the LET of the proton beam. Part of the hesitation to incorporate variable RBE models in the clinic is due to the great noise in the biological datasets on which these models are trained, often making it unclear which model, if any, provides sufficiently accurate RBE predictions to warrant a departure from RBE = 1.1. PURPOSE: Here, we introduce a novel model of proton RBE based on how a cell's intrinsic radiosensitivity varies with LET, rather than its LQM parameters. METHODS AND MATERIALS: We performed clonogenic cell survival assays for eight cell lines exposed to 6 MV x-rays and 1.2, 2.6, or 9.9 keV/µm protons, and combined our measurements with published survival data (n = 397 total cell line/LET combinations). We characterized how radiosensitivity metrics of the form DSF% , (the dose required to achieve survival fraction [SF], e.g., D10% ) varied with proton LET, and calculated the Bayesian information criteria associated with different LET-dependent functions to determine which functions best described the underlying trends. This allowed us to construct a six-parameter model that predicts cells' proton survival curves based on the LET dependence of their radiosensitivity, rather than the LET dependence of the LQM parameters themselves. We compared the accuracy of our model to previously established empirical proton RBE models, and implemented our model within a clinical treatment plan evaluation workflow to demonstrate its feasibility in a clinical setting. RESULTS: Our analyses of the trends in the data show that DSF% is linearly correlated between x-rays and protons, regardless of the choice of the survival level (e.g., D10% , D37% , or D50% are similarly correlated), and that the slope and intercept of these correlations vary with proton LET. The model we constructed based on these trends predicts proton RBE within 15%-30% at the 68.3% confidence level and offers a more accurate general description of the experimental data than previously published empirical models. In the context of a clinical treatment plan, our model generally predicted higher RBE-weighted doses than the other empirical models, with RBE-weighted doses in the distal portion of the field being up to 50.7% higher than the planned RBE-weighted doses (RBE = 1.1) to the tumor. CONCLUSIONS: We established a new empirical proton RBE model that is more accurate than previous empirical models, and that predicts much higher RBE values in the distal edge of clinical proton beams.

Effect of boron compounds on the biological effectiveness of proton therapy.

PURPOSE: We assessed whether adding sodium borocaptate (BSH) or 4-borono-l-phenylalanine (BPA) to cells irradiated with proton beams influenced the biological effectiveness of those beams against prostate cancer cells to investigate if the alpha particles generated through proton-boron nuclear reactions would be sufficient to enhance the biological effectiveness of the proton beams. METHODS: We measured clonogenic survival in DU145 cells treated with 80.4-ppm BSH or 86.9-ppm BPA, or their respective vehicles, after irradiation with 6-MV X-rays, 1.2-keV/µm (low linear energy transfer [LET]) protons, or 9.9-keV/µm (high-LET) protons. We also measured γH2AX and 53BP1 foci in treated cells at 1 and 24 h after irradiation with the same conditions. RESULTS: We found that BSH radiosensitized DU145 cells across all radiation types. However, no difference was found in relative radiosensitization, characterized by the sensitization enhancement ratio or the relative biological effectiveness, for vehicle- versus BSH-treated cells. No differences were found in numbers of γH2AX or 53BP1 foci or γH2AX/53BP1 colocalized foci for vehicle- versus BSH-treated cells across radiation types. BPA did not radiosensitize DU145 cells nor induced any significant differences when comparing vehicle- versus BPA-treated cells for clonogenic cell survival or γH2AX and 53BP1 foci or γH2AX/53BP1 colocalized foci. CONCLUSIONS: Treatment with 11 B, at concentrations of 80.4 ppm from BSH or 86.9 ppm from BPA, had no effect on the biological effectiveness of proton beams in DU145 prostate cancer cells. Our results agree with published theoretical calculations indicating that the contribution of alpha particles from such reactions to the total absorbed dose and biological effectiveness is negligible. We also found that BSH radiosensitized DU145 cells to X-rays, low-LET protons, and high-LET protons but that the radiosensitization was not related to DNA damage.

Cell lines of the same anatomic site and histologic type show large variability in intrinsic radiosensitivity and relative biological effectiveness to protons and carbon ions.

PURPOSE: To show that intrinsic radiosensitivity varies greatly for protons and carbon (C) ions in addition to photons, and that DNA repair capacity remains important in governing this variability. METHODS: We measured or obtained from the literature clonogenic survival data for a number of human cancer cell lines exposed to photons, protons (9.9 keV/µm), and C-ions (13.3-77.1 keV/µm). We characterized their intrinsic radiosensitivity by the dose for 10% or 50% survival (D10% or D50% ), and quantified the variability at each radiation quality by the coefficient of variation (COV) in D10% and D50% . We also treated cells with DNA repair inhibitors prior to irradiation to assess how DNA repair capacity affects their variability. RESULTS: We found no statistically significant differences in the COVs of D10% or D50% between any of the radiation qualities investigated. The same was true regardless of whether the cells were treated with DNA repair inhibitors, or whether they were stratified into histologic subsets. Even within histologic subsets, we found remarkable differences in radiosensitivity for high LET C-ions that were often greater than the variations in RBE, with brain cancer cells varying in D10% (D50% ) up to 100% (131%) for 77.1 keV/µm C-ions, and non-small cell lung cancer and pancreatic cancer cell lines varying up to 55% (76%) and 51% (78%), respectively, for 60.5 keV/µm C-ions. The cell lines with modulated DNA repair capacity had greater variability in intrinsic radiosensitivity across all radiation qualities. CONCLUSIONS: Even for cell lines of the same histologic type, there are remarkable variations in intrinsic radiosensitivity, and these variations do not differ significantly between photon, proton or C-ion radiation. The importance of DNA repair capacity in governing the variability in intrinsic radiosensitivity is not significantly diminished for higher LET radiation.

Translational pharmacology of an inhaled small molecule αvß6 integrin inhibitor for idiopathic pulmonary fibrosis.

The αvß6 integrin plays a key role in the activation of transforming growth factor-ß (TGFß), a pro-fibrotic mediator that is pivotal to the development of idiopathic pulmonary fibrosis (IPF). We identified a selective small molecule αvß6 RGD-mimetic, GSK3008348, and profiled it in a range of disease relevant pre-clinical systems. To understand the relationship between target engagement and inhibition of fibrosis, we measured pharmacodynamic and disease-related end points. Here, we report, GSK3008348 binds to αvß6 with high affinity in human IPF lung and reduces downstream pro-fibrotic TGFß signaling to normal levels. In human lung epithelial cells, GSK3008348 induces rapid internalization and lysosomal degradation of the αvß6 integrin. In the murine bleomycin-induced lung fibrosis model, GSK3008348 engages αvß6, induces prolonged inhibition of TGFß signaling and reduces lung collagen deposition and serum C3M, a marker of IPF disease progression. These studies highlight the potential of inhaled GSK3008348 as an anti-fibrotic therapy.

Isolated Congenital Round Window Atresia: Report of 2 Cases.

Round window atresia (RWA) is an uncommon condition and can result in a conductive hearing loss. Two cases of nonsyndromal bilateral RWA in 2 members of the same family are reported. Both cases presented with a conductive hearing loss of 20 to 30 dB. High-resolution computed tomography scanning was used to diagnose the condition. The patients were rehabilitated with hearing aids. Review of the literature has shown disappointing results in hearing improvement with cochlear fenestration in an attempt to address this condition. Patients presenting with unexplained conductive hearing loss should be offered computed tomography scanning. The cases we report add to the literature to benefit future patients in preoperative counseling and better inform management.

Isolation of time-dependent DNA damage induced by energetic carbon ions and their fragments using fluorescent nuclear track detectors.

PURPOSE: High energetic carbon (C-) ion beams undergo nuclear interactions with tissue, producing secondary nuclear fragments. Thus, at depth, C-ion beams are composed of a mixture of different particles with different linear energy transfer (LET) values. We developed a technique to enable isolation of DNA damage response (DDR) in mixed radiation fields using beam line microscopy coupled with fluorescence nuclear track detectors (FNTDs). METHODS: We imaged live cells on a coverslip made of FNTDs right after C-ion, proton or photon irradiation using an in-house built confocal microscope placed in the beam path. We used the FNTD to link track traversals with DNA damage and separated DNA damage induced by primary particles from fragments. RESULTS: We were able to spatially link physical parameters of radiation tracks to DDR in live cells to investigate spatiotemporal DDR in multi-ion radiation fields in real time, which was previously not possible. We demonstrated that the response of lesions produced by the high-LET primary particles associates most strongly with cell death in a multi-LET radiation field, and that this association is not seen when analyzing radiation induced foci in aggregate without primary/fragment classification. CONCLUSIONS: We report a new method that uses confocal microscopy in combination with FNTDs to provide submicrometer spatial-resolution measurements of radiation tracks in live cells. Our method facilitates expansion of the radiation-induced DDR research because it can be used in any particle beam line including particle therapy beam lines. CATEGORY: Biological Physics and Response Prediction.

Nonhomologous End Joining Is More Important Than Proton Linear Energy Transfer in Dictating Cell Death.

PURPOSE: This study seeks to identify biological factors that may yield a therapeutic advantage of proton therapy versus photon therapy. Specifically, we address the role of nonhomologous end-joining (NHEJ) and homologous recombination (HR) in the survival of cells in response to clinical photon and proton beams. METHODS AND MATERIALS: We irradiated HT1080, M059K (DNA-PKcs+/+), and HCC1937 human cancer cell lines and their isogenic counterparts HT1080-shDNA-PKcs, HT1080-shRAD51IND, M059J (DNA-PKcs-/-), and HCC1937-BRCA1 (BRCA1 complemented) to assess cell clonogenic survival and γ-H2AX radiation-induced foci. Cells were irradiated with either clinically relevant photons or 1 of 3 proton linear energy transfer (LET) values. RESULTS: Our results indicate that NHEJ deficiency is more important in dictating cell survival than proton LET. Cells with disrupted HR through BRCA1 mutation showed increased radiosensitivity only for high-LET protons whereas RAD51 depletion showed increased radiosensitivity for both photons and protons. DNA double strand breaks, assessed by γ-H2AX radiation-induced foci, showed greater numbers after 24 hours in cells exposed to higher LET protons. We also observed that NHEJ-deficient cells were unable to repair the vast majority of double strand breaks after 24 hours. CONCLUSIONS: BRCA1 mutation significantly sensitizes cells to protons, but not photons. Loss of NHEJ renders cells hypersensitive to radiation, whereas the relative importance of HR increases with LET across several cell lines. This may be attributable to the more clustered damage induced by higher LET protons, which are harder to repair through NHEJ. This highlights the importance of tumor biology in dictating treatment modality and suggests BRCA1 as a potential biomarker for proton therapy response. Our data also support the use of pharmacologic inhibitors of DNA repair to enhance the sensitivity to different radiation types, although this raises issues for normal tissue toxicity.

Nasal microbial composition and chronic otitis media with effusion: A case-control study.

OBJECTIVES: Chronic otitis media with effusion (COME) in children can cause prolonged hearing loss, which is associated with an increased risk of learning delays and behavioural problems. Dispersal of bacterial pathogens from the nasal passages to the middle ear is implicated in COME. We sought to determine whether there is an association between nasal microbial composition and COME in children. METHODS: A case-control study of children aged 3 and 4 years was conducted. Cases undergoing placement of tympanostomy tubes for COME were compared to healthy controls. Nasal swabs were collected and a questionnaire was administered. The V1-3 region of the 16S rRNA gene was amplified, and sequenced on the Illumina MiSeq. RESULTS: 73 children with COME had a lower Shannon diversity index than 105 healthy controls (1.62 [.80] versus 1.88 [.84], respectively; P = .046). The nasal microbiota of cases and controls differed in composition using Bray-Curtis dissimilarity (p = 0.002). Children with COME had a higher abundance of otopathogens and lower abundance of commensals including alpha haemolytic Streptococci and Lactococcus. Cluster analysis revealed 4 distinct nasal microbial profiles. Profiles that were Corynebacterium-dominated (aOR 4.18 [95%CI, 1.68-10.39], Streptococcus-dominated (aOR 3.12 [95%CI, 1.08-9.06], or Moraxella-dominated (aOR 4.70 [95%CI, 1.73-12.80] were associated with COME, compared to a more mixed microbial profile when controlling for age, ethnicity, and recent antibiotics use. CONCLUSIONS: Children with COME have a less diverse nasal microbial composition with a higher abundance of pathogens, compared to healthy children who have a more mixed bacterial profile with a higher abundance of commensals. Further research is required to determine how nasal microbiota may relate to the pathogenesis or maintenance of COME, and whether modification of the nasal microbiota can prevent or treat children at risk of COME.

Drug screening of biopsy-derived spheroids using a self-generated microfluidic concentration gradient.

Performing drug screening of tissue derived from cancer patient biopsies using physiologically relevant 3D tumour models presents challenges due to the limited amount of available cell material. Here, we present a microfluidic platform that enables drug screening of cancer cell-enriched multicellular spheroids derived from tumour biopsies, allowing extensive anticancer compound screening prior to treatment. This technology was validated using cell lines and then used to screen primary human prostate cancer cells, grown in 3D as a heterogeneous culture from biopsy-derived tissue. The technology enabled the formation of repeatable drug concentration gradients across an array of spheroids without external fluid actuation, delivering simultaneously a range of drug concentrations to multiple sized spheroids, as well as replicates for each concentration. As proof-of-concept screening, spheroids were generated from two patient biopsies and a panel of standard-of-care compounds for prostate cancer were tested. Brightfield and fluorescence images were analysed to provide readouts of spheroid growth and health, as well as drug efficacy over time. Overall, this technology could prove a useful tool for personalised medicine and future drug development, with the potential to provide cost- and time-reduction in the healthcare delivery.

Higher serum 25(OH)D concentration is associated with lower risk of chronic otitis media with effusion: a case-control study.

AIM: Vitamin D supplementation and higher 25(OH)-vitamin D concentration are associated with reduced risk of acute respiratory infection. This study examined whether there is a similar association between higher serum 25(OH)D concentration and lower risk of chronic otitis media with effusion (COME). METHODS: In a case-control study, serum 25(OH)D concentration in children referred for tympanostomy tube placement for COME (n = 178) was compared to that of healthy children randomly sampled from primary care practices (n = 179). Subjects aged three and four years were recruited in Auckland, New Zealand between May 2011 and November 2013. Blood samples were collected from the children, and their guardians were interviewed. Odds ratios were calculated using logistic regression. RESULTS: In a multivariable analysis, higher serum 25(OH)D concentration was associated with a lower risk of COME (OR: 0.86 per 10 nmol/L; 95% CI 0.77-0.97) after adjusting for age, sex, deprivation index, ethnicity, tobacco smoke exposure, duration of breastfeeding and season of blood sampling. Further adjustment for eight additional risk factors did not change the result. CONCLUSION: This finding supports further investigation into whether the risk of COME could be reduced by increasing serum 25(OH)D concentration through increased sun exposure, higher dietary intake or vitamin D supplementation.

Determinants of chronic otitis media with effusion in preschool children: a case-control study.

BACKGROUND: Chronic otitis media with effusion (COME) is a prevalent upper airway infection resulting in hearing loss. The aim of this research was to determine risk factors for COME in preschool children. METHODS: A case-control design was conducted in Auckland, New Zealand from May 2011 until November 2013. The cases were children aged 3 and 4 years referred for tympanostomy tube placement due to a diagnosis of COME (n = 178). The controls were a random sample of healthy children aged 3 and 4 years from primary care practices (n = 209). The children's guardians completed an interviewer-administered questionnaire that covered topics including socio-demographic information, pregnancy and birth, infant feeding practices, home environment, and respiratory health. In addition, skin prick tests for atopy were performed. Odds ratios (OR) estimating the risk of COME independently associated with the exposures were calculated using a logistic regression model. RESULTS: Children with COME frequently had nasal obstruction (OR: 4.38 [95% CI: 2.37-8.28]), always snored (OR: 3.64 [95% CI: 1.51-9.15]) or often snored (OR: 2.45 [95% CI: 1.04-5.96]), spent more hours per week in daycare (OR per hour/week: 1.03 [95% CI: 1.00-1.05]), had frequent colds (OR: 2.67 [95% CI: 1.59-4.53]), had siblings who had undergone tympanostomy tube placement (OR: 2.68 [95% CI: 1.22-6.02]), underwent long labour (OR: 2.59 [95% CI: 1.03-6.79]), and had early introduction of cow's milk (OR: 1.76 [95% CI: 1.05-2.97]). Asian ethnicity (OR: 0.20 [95% CI: 0.07-0.53]) and having older siblings (OR: 0.54 [95% CI: 0.31-0.93]) were inversely associated with COME. CONCLUSION: COME in preschool children was associated with pathogen exposure, respiratory infection, and nasal obstruction. Strategies to prevent pathogen transmission warrant investigation. The novel findings of long labour and early cow's milk introduction require replication in future studies.

Time-Lapse Monitoring of DNA Damage Colocalized With Particle Tracks in Single Living Cells.

PURPOSE: Understanding the DNA damage and repair induced by hadron therapy (HT) beams is crucial for developing novel strategies to maximize the use of HT beams to treat cancer patients. However, spatiotemporal studies of DNA damage and repair for beam energies relevant to HT have been challenging. We report a technique that enables spatiotemporal measurement of radiation-induced damage in live cells and colocalization of this damage with charged particle tracks over a broad range of clinically relevant beam energies. The technique uses novel fluorescence nuclear track detectors with fluorescence confocal laser scanning microscopy in the beam line to visualize particle track traversals within the subcellular compartments of live cells within seconds after injury. METHODS AND MATERIALS: We designed and built a portable fluorescence confocal laser scanning microscope for use in the beam path, coated fluorescence nuclear track detectors with fluorescent-tagged live cells (HT1080 expressing enhanced green fluorescent protein tagged to XRCC1, a single-strand break repair protein), placed the entire assembly into a proton therapy beam line, and irradiated the cells with a fluence of ∼1 × 10(6) protons/cm(2). RESULTS: We successfully obtained confocal images of proton tracks and foci of DNA single-strand breaks immediately after irradiation. CONCLUSIONS: This technique represents an innovative method for analyzing biological responses in any HT beam line at energies and dose rates relevant to therapy. It allows precise determination of the number of tracks traversing a subcellular compartment and monitoring the cellular damage therein, and has the potential to measure the linear energy transfer of each track from therapeutic beams.

Pharmacological Characterization of the αvß6 Integrin Binding and Internalization Kinetics of the Foot-and-Mouth Disease Virus Derived Peptide A20FMDV2.

A20FMDV2 is a peptide derived from the foot-and-mouth disease virus with a high affinity and selectivity for the alpha-v beta-6 (αvß6) arginyl-glycinyl-aspartic acid (RGD)-binding integrin. It has been shown to be an informative tool ligand in pre-clinical imaging studies for selective labelling of the αvß6 integrin in a number of disease models. In a radioligand binding assay using a radiolabelled form of the peptide ([3H]A20FMDV2), its high affinity (K(D): 0.22 nmol/l) and selectivity (at least 85-fold) for αvß6 over the other members of the RGD integrin family was confirmed. [3H]A20FMDV2 αvß6 binding could be fully reversed only in the presence of EDTA, whereas a partial reversal was observed in the presence of excess concentrations of an RGD-mimetic small molecule (SC-68448) or unlabelled A20FMDV2. Using flow cytometry on bronchial epithelial cells, the ligand-induced internalization of αvß6 by A20FMDV2 and latency-associated peptide-1 was shown to be fast (t(1/2): 1.5 and 3.1 min, respectively), concentration-dependent (EC50: values 1.1 and 3.6 nmol/l, respectively) and was followed by a moderately slow return of integrin to the surface. The results of the radioligand binding studies suggest that the binding of A20FMDV2 to the RGD-binding site on αvß6 is required to maintain its engagement with the hypothesised A20FMDV2 synergy site on the integrin. In addition, there is evidence from flow cytometric studies that the RGD-ligand engagement of αvß6 post-internalization plays a role in delaying recycling of the integrin to the cell surface. This mechanism may act as a homeostatic control of membrane αvß6 following RGD ligand engagement.

Post-weaning increases in the milk-fat globule EGF-factor VIII on fat globules in mouse milk and in the uptake of the fat globules by HC11 mammary epithelial cells.

Milk fat globules (MFGs) secreted by lactating mammary gland are unique lipid surrounded by a phospholipid bi-layer. We report here post-weaning changes in MFG EGF factor VIII (MFG-E8) and annexin V-accessible phosphatidyl-l-serine on the surface of MFGs. The MFG content in milk markedly decreased to about one-half within 2 days after forced weaning, despite a slight increase in milk protein content. Immunofluorescence-staining of MFGs using anti-MFG-E8 and annexin V indicated that MFG-E8 was present on some, but not all, MFGs before weaning, whereas most of MFGs were MFG-E8-positive and annexin V-negative after weaning. Free MFG-E8 with binding activity to phosphatidyl-l-serine was present abundantly in the post-weaning milk, and indeed exhibited binding to MFGs in pre-weaning milk. MFGs were taken up by HC11 mouse mammary epithelial cells in vitro, and those from post-weaning milk were remarkable for such cellular uptake. Moreover, the uptake of MFGs by the cells was inhibited by an anti-MFG-E8 antibody. Taken together, these findings suggest that MFG-E8 plays a critical role in regulation of MFG dynamics after weaning or during the suckling interval through the control of MFG-epithelial cell interaction in lactating mammary glands.

Cell motility in models of wounded human skin is improved by Gap27 despite raised glucose, insulin and IGFBP-5.

Reducing Cx43 expression stimulates skin wound healing. This is mimicked in models when Cx43 function is blocked by the connexin mimetic peptide Gap27. IGF-I also stimulates wound healing with IGFBP-5 attenuating its actions. Further, the IGF-I to IGFBP-5 ratio is altered in diabetic skin, where wound closure is impaired. We investigated whether Gap27 remains effective in augmenting scrape-wound closure in human skin wound models simulating diabetes-induced changes, using culture conditions with raised glucose, insulin and IGFBP-5. Gap27 increased scrape-wound closure in normal glucose and insulin (NGI) and to a lesser extent in high glucose and insulin (HGI). IGF-I enhanced scrape-wound closure in keratinocytes whereas IGFBP-5 inhibited this response. Gap27 overcame the inhibitory effects of IGFBP-5 on IGF-I activity. Connexin-mediated communication (CMC) was reduced in HGI, despite raised Cx43, and Gap27 significantly decreased CMC in NGI and HGI. IGF-I and IGFBP-5 did not affect CMC. IGF-I increased keratinocyte proliferation in NGI, and Gap27 increased proliferation in NGI to a greater extent than in HGI. We conclude that IGF-I and Gap27 stimulate scrape-wound closure by independent mechanisms with Gap27 inhibiting Cx43 function. Gap27 can enhance wound closure in diabetic conditions, irrespective of the IGF-I:IGFBP-5 balance.

Myofibroma of the mandible: a case report.

This report describes the case of a male aged 28 years who presented with a chief complaint of discomfort and swelling in the mandibular right molar area. An incisional biopsy was performed with a preliminary differential diagnosis of periodontal abscess, fibrotic lesion, or odontogenic tumor. Subsequent excision of the lesion was performed and histologic analysis confirmed the diagnosis of myofibroma.

IGFBP5 induces cell adhesion, increases cell survival and inhibits cell migration in MCF-7 human breast cancer cells.

Maintenance of tissue boundaries is crucial for control of metastasis. We describe a new signalling pathway in which epithelial cell disruption can be minimised and thereby restricts epithelial-mesenchymal transgressions. This involves the release of insulin-like growth factor (IGF)-binding protein 5 (IGFBP5) from apoptotic cells, which increases the adhesion of epithelial cells on mesenchymal but not epithelial extracellular matrix (ECM), and involves the direct interaction of IGFBP5 and α2ß1 integrins. IGFBP5 also induced cell adhesion to vitronectin in the absence of αVß3 integrin, the vitronectin receptor, again through an α2ß1-integrin-dependent action, suggesting that IGFBP5 can induce spreading on matrices, even in the absence of the integrins normally used in this process. Using IGFBP5 mutants we demonstrate that the effect is IGF-independent but requires the heparin-binding domain in the C-terminus of IGFBP5. A truncated mutant containing only the C-terminal of IGFBP5 also induced adhesion. Adhesion induced by IGFBP5 was dependent on Cdc42 and resulted in activation of integrin-linked kinase (ILK) and Akt. Consistent with these changes, IGFBP5 facilitated prolonged cell survival in nutrient-poor conditions and decreased phosphorylation of the stress-activated kinase p38 MAPK (MAPK14). Whereas IGFBP5 enhanced adhesion, it inhibited cell migration, although this was not evident using the truncated C-terminal mutant, suggesting that effects of IGFBP5 on adhesion and migration involve different mechanisms. We anticipate that these responses to IGFBP5 would reduce the metastatic potential of cells.