Measurement Properties of the Canadian Occupational Performance Measure: A Systematic Review.

IMPORTANCE: The Canadian Occupational Performance Measure (COPM) is widely used in clinical practice and research. However, the measurement properties of the COPM were not reviewed using rigorous systematic methodology. OBJECTIVE: To evaluate the measurement properties of the COPM. DATA SOURCES: MEDLINE, Web of Science, Scopus, OTseeker, and Cochrane Library. Study Selection and Data Collection: We used the updated COnsensus-based Standards for the selection of health Measurement INstruments (COSMIN) Risk of Bias checklist to evaluate the measurement properties of the COPM reported in relevant studies. FINDINGS: Our search identified 35 articles that reported measurement properties for the COPM with samples that differed in age, country, diagnosis, and disease stage. For content validity, the evidence was inconsistent and of low quality; no studies assessed structural validity. For reliability, the internal consistency was indeterminate and of low quality. One study reported indeterminate and very low quality evidence for cross-cultural validity. According to the evidence reported in these studies, the COPM has inconsistent and moderate reliability, construct validity, and responsiveness and insufficient and high-quality evidence for criterion validity. CONCLUSIONS AND RELEVANCE: Our review of the evidence using the COSMIN Risk of Bias checklist indicates that the Canadian Occupational Performance Measure lacks high-quality validation. What This Article Adds: High-quality validation of the Canadian Occupational Performance Measure is lacking. Further examination of its measurement properties using updated relevant guidelines is required.

The effect on subjective quality of life of occupational therapy based on adjusting the challenge-skill balance: a randomized controlled trial.

OBJECTIVE: To verify the effect of adjusting the challenge-skill balance with respect to rehabilitation process. DESIGN: A single-blind, two-arm, parallel-group, randomized controlled trial. SETTING: Recovery rehabilitation unit of Harue Hospital, Japan. SUBJECTS: The trial included 72 clients (mean (SD): age, 74.64 (9.51) years; Functional Independence Measure score, 98.26 (15.27)) with cerebral or spinal disease or musculoskeletal disease. INTERVENTIONS: Clients were randomly divided into two groups: the experimental group, who received occupational therapy with adjustment of the challenge-skill balance, and the control group who received conventional occupational therapy. Time from admission to discharge was considered the implementation period; the final evaluation was conducted at three months after discharge. MAIN MEASURES: The primary outcome was subjective quality of life (Ikigai-9). Secondary outcomes were the health-related quality of life (EuroQol-5 Dimensions, Five Levels (EQ-5D-5L)), the Flow State Scale for Occupational Tasks, and the Functional Independence Measure. A cost-effectiveness analysis was conducted using total cost and quality-adjusted life-year based on the EQ-5D-5L. RESULTS: Significant differences were observed between the experimental and control groups with respect to the Ikigai-9 score (P = 0.008) and EQ-5D-5L (P = 0.038), and the effect sizes were 0.76 (95% confidence interval [CI]: 0.27-1.24) and 0.62 (95% CI: 0.14-1.10), respectively. No significant between-group differences in other outcomes were observed, for example, the Functional Independence Measure score improved in both experimental and control groups (119.80 (5.50) and 118.84 (6.97), respectively. The incremental cost-effectiveness ratio was US$5518.38. CONCLUSIONS: Adjusting the challenge-skill balance may be a useful approach to improve the participant's subjective quality of life in the rehabilitation process.

CUL4-DDB1 ubiquitin ligase interacts with multiple WD40-repeat proteins and regulates histone methylation.

The CUL4-DDB1-ROC1 ubiquitin E3 ligase regulates cell-cycle progression, replication and DNA damage response. However, the substrate-specific adaptors of this ligase remain uncharacterized. Here, we show that CUL4-DDB1 complexes interact with multiple WD40-repeat proteins (WDRs) including TLE1-3, WDR5, L2DTL (also known as CDT2) and the Polycomb-group protein EED (also known as ESC). WDR5 and EED are core components of histone methylation complexes that are essential for histone H3 methylation and epigenetic control at K4 or K9 and K27, respectively, whereas L2DTL regulates CDT1 proteolysis after DNA damage through CUL4-DDB1 (ref. 8). We found that CUL4A-DDB1 interacts with H3 methylated mononucleosomes and peptides. Inactivation of either CUL4 or DDB1 impairs these histone modifications. However, loss of WDR5 specifically affects histone H3 methylation at K4 but not CDT1 degradation, whereas inactivation of L2DTL prevents CDT1 degradation but not histone methylation. Our studies suggest that CUL4-DDB1 ligases use WDR proteins as molecular adaptors for substrate recognition, and modulate multiple biological processes through ubiquitin-dependent proteolysis.

Tyrosine phosphorylation controls PCNA function through protein stability.

The proliferating cell nuclear antigen (PCNA) is an essential protein for DNA replication and damage repair. How its function is controlled remains an important question. Here, we show that the chromatin-bound PCNA protein is phosphorylated on Tyr 211, which is required for maintaining its function on chromatin and is dependent on the tyrosine kinase activity of EGF receptor (EGFR) in the nucleus. Phosphorylation on Tyr 211 by EGFR stabilizes chromatin-bound PCNA protein and associated functions. Consistently, increased PCNA Tyr 211 phosphorylation coincides with pronounced cell proliferation, and is better correlated with poor survival of breast cancer patients, as well as nuclear EGFR in tumours, than is the total PCNA level. These results identify a novel nuclear mechanism linking tyrosine kinase receptor function with the regulation of the PCNA sliding clamp.

Direct roles of the signaling kinase RSK2 in Cdc25C activation during Xenopus oocyte maturation.

The induction of M phase in eukaryotic cell cycles requires robust activation of Cdc2/cyclin B by Cdc25, which itself is robustly activated by serine/threonine phosphorylations. Although multiple protein kinases that directly activate Cdc25C have been identified, whether the combination of different primary phosphorylations of Cdc25C is sufficient to fully activate Cdc25C has not been determined. By analyzing the GST-Cdc25C phosphorylating activity in Xenopus egg extracts, we previously defined roles of MAPK and Cdc2/cyclin B in partially activating Cdc25C and predicted the presence of another major Cdc25C-activating kinase. In this study, we demonstrate that this missing kinase is RSK2, which phosphorylates three sites in Cdc25C and also partially activates Cdc25C. However, the phosphorylations catalyzed by MAPK, Cdc2, and RSK2 fail to fully activate Cdc25C, suggesting that additional biochemical events are required to fully activate this key cell cycle regulator.

Aberrant expression of katanin p60 in prostate cancer bone metastasis.

BACKGROUND: Katanin p60 is a microtubule-severing protein and is involved in microtubule cytoskeleton organization in both mitotic and non-mitotic processes. Its role in cancer metastasis is unknown. METHODS: Differential protein profiles of bone marrow aspirates were analyzed by chromatography, electrophoresis, and mass spectrometry. Expression of katanin p60 in primary and metastatic prostate cancer was examined by immunohistochemistry. Cellular function of katanin p60 was further examined in prostate cell lines. RESULTS: In a proteomic profiling of bone marrow aspirates from men with prostate cancer, we found that katanin p60 was one of the proteins differentially expressed in bone metastasis samples. Immunohistochemical staining showed that katanin p60 was expressed in the basal cells in normal human prostate glands. In prostatic adenocarcinomas, in which the basal cells were absent, katanin p60 was expressed in the prostate cancer cells. In the specimens from bone metastasis, katanin p60 was detectable in the metastatic cancer cells. Strikingly, some of the metastatic cancer cells also co-expressed basal cell biomarkers including the tumor suppressor p53-homologous protein p63 and the high molecular weight cytokeratins, suggesting that the metastatic prostate cancer cells may have a basal cell-like phenotype. Moreover, overexpression of katanin p60 inhibited prostate cancer cell proliferation but enhanced cell migration activity. CONCLUSIONS: Katanin p60 was aberrantly expressed during prostate cancer progression. Its expression in the metastatic cells in bone was associated with the re-emergence of a basal cell-like phenotype. The elevated katanin p60 expression may contribute to cancer cell metastasis via a stimulatory effect on cell motility.

Ubiquitin ligase component Cul4 associates with Clr4 histone methyltransferase to assemble heterochromatin.

In eukaryotes, heterochromatin mediates diverse processes including gene silencing and regulation of long-range chromatin interactions. The formation of heterochromatin involves a conserved array of histone modifications; in particular, methylation of histone H3 at Lys 9 (H3K9me) is essential for recruiting HP1/Swi6 proteins. In fission yeast, the Clr4 methyltransferase is responsible for H3K9me across all heterochromatic domains. However, the mechanism of Clr4 recruitment to these loci is poorly understood. We show that Clr4 associates with Cul4, a cullin family protein that serves as a scaffold for assembling ubiquitin ligases. Mutations in Cul4 result in defective localization of Clr4 and loss of silencing at heterochromatic loci. This is accompanied by a severe reduction in H3K9me and Swi6 levels, and accumulation of transcripts corresponding to naturally silenced repeat elements within heterochromatic domains. Moreover, heterochromatin defects in Cul4 mutants could not be rescued by expression of Cul4 protein lacking Nedd8 modification, which is essential for its ubiquitin ligase activity. Rik1, a protein related to DNA damage binding protein DDB1 and required for H3K9me, also interacts with Cul4, the association of which might serve to target Clr4 to heterochromatic loci. These analyses uncover a role for Cul4-based protein ubiquitination in regulating H3K9me and heterochromatin formation.

Distinct roles of HDAC complexes in promoter silencing, antisense suppression and DNA damage protection.

Histone acetylation is important in regulating DNA accessibility. Multifunctional Sin3 proteins bind histone deacetylases (HDACs) to assemble silencing complexes that selectively target chromatin. We show that, in fission yeast, an essential HDAC, Clr6, exists in two distinct Sin3 core complexes. Complex I contains an essential Sin3 homolog, Pst1, and other factors, and predominantly targets gene promoters. Complex II contains a nonessential Sin3 homolog, Pst2, and several conserved proteins. It preferentially targets transcribed chromosomal regions and centromere cores. Defects in complex II abrogate global protective functions of chromatin, causing increased accessibility of DNA to genotoxic agents and widespread antisense transcripts that are processed by the exosome. Notably, the two Clr6 complexes differentially repress forward and reverse centromeric repeat transcripts, suggesting that these complexes regulate transcription in heterochromatin and euchromatin in similar manners, including suppression of spurious transcripts from cryptic start sites.

Flow experience and health-related quality of life in community dwelling elderly Japanese.

This study was performed to investigate the relationship between mental state (especially flow experience [Flow Experience Checklist of Ishimura]) health-related quality of life (Medical Outcome Study 8-Item Short-Form Health Survey), and stress (salivary amylase concentrations). The subjects were 119 healthy elderly persons living in rural areas who participated in a "meeting for the elderly" at a nursing home in Kagawa Prefecture, Japan. The subjects were classified into apathetic, anxious, relaxed, and flow groups. It was found that physical health was significantly better when performing important daily activities in the group who experienced flow and the relaxed group than in the group that was in an apathetic state. However, no significant relationship was observed between the degree of flow experience and stress. The present findings suggested that interventions which make the activities of daily life either "high-challenge, high-skill" situations or "low-challenge, high-skill" situations could have a positive influence on the physical health of community-living elderly Japanese. A longitudinal study should be performed in the future.

A Pilot Study of a Group Program Focused on Enabling Life Performance for Older Adults Living in the Community.

Background: This study attempts to determine whether a program focused on improving literacy in daily living is effective in preventing physical frailty, and to compare standard treatments for physical frailty. Methods: This study was designed as a pilot intervention study involving two groups. Twenty-five older adults aged 65 to 85 in Ward A, Tokyo, were randomly assigned to the literacy group or the exercise group on a regional basis and were given a 60- to 90-minute program twice a month, eight times over four months. The literacy group mainly used video materials to monitor learning, and the exercise group used a multifactor exercise program. Results: The LSI-Z, GAS-L, Maximum 5 m walking time, and TUG tests showed the main effects before and after the intervention in both groups (p < 0.05, p < 0.01). The WHOQOL26, Maximum 5 m walking time, and TUG tests also showed the main effects across both groups (p < 0.05). Conclusion: Both programs, when implemented independently, showed specific effects on subjective well-being, occupational performance, and physical fitness. However, QOL and physical fitness were significantly higher in the exercise group than in the literacy group. These results should be considered with caution because of the limited sample size of this pilot study.

Reliability and Validity of the Japanese Version of the Assessment of Readiness for Mobility Transition (ARMT-J) for Japanese Elderly.

The Assessment of Readiness for Mobility Transition (ARMT) questionnaire assesses individuals' emotional and attitudinal readiness related to mobility as they age. This study aimed to examine the reliability and validity of the Japanese version of the ARMT (ARMT-J). The ARMT-J and related variables were administered to 173 patients and staff members undergoing rehabilitation at hospitals in Japan. Construct validity was first examined using confirmatory factor analysis (CFA) to confirm cross-cultural validity. For structural validity, the optimal number of factors was confirmed using a Velicer's minimum average partial test and parallel analysis, followed by exploratory factor analysis (EFA). Finally, a CFA was performed using the most appropriate model. Internal consistency, test-retest reliability, standard error of measurement (SEM), and smallest detectable change (SDC) were assessed for reliability. The CFA fit for the factor structure of the original ARMT was low. Therefore, the EFA was conducted with two to four factors. The optimal factor structure was three factors, with a Cronbach's alpha coefficient and Cohen's weighted kappa coefficient of 0.85 and 0.76, respectively. The intraclass correlation coefficient (ICC) of the test-retest was 0.93, the SEM was 0.72, and the SDC was 2.00. The model fit was good for the ARMT-J, with a three-factor structure.

Developing an international research of health-care ICT applied for rehabilitation and daily living support between Japan and the Netherlands.

While many health-care issues and technological solutions are viewed locally, developing new technological solutions might benefit from lessons learned globally. The aim of this study was to develop a shared international research agenda of health-care ICT, applied to rehabilitation and daily living support. This study was focused on sensor technology and social robots used for supporting older persons in the Netherlands (Amsterdam) and Japan (Tokyo). Three researchers from Amsterdam University of Applied Sciences visited Japan and four researchers from Tokyo Metropolitan University visited the Netherlands and conducted field-visits and mutual presentations. Using a nominal group technique (NGT) facilitated the expert panel deliberations. Research priorities were identified qualitatively through in-action critical reflection on emerging ideas, and quantitatively by ranking of identified knowledge gaps (using the Mentimeter© app). The resulting joint research agenda identified topics around the utility of sensor monitoring and processes of acceptance of health-care ICT among older persons and occupational therapists. The agenda was complemented by formulating underlying assumptions prescribing such research to be embedded in real-life situations with the participation of stakeholders.

A role for cell-cycle-regulated histone H3 lysine 56 acetylation in the DNA damage response.

DNA breaks are extremely harmful lesions that need to be repaired efficiently throughout the genome. However, the packaging of DNA into nucleosomes is a significant barrier to DNA repair, and the mechanisms of repair in the context of chromatin are poorly understood. Here we show that lysine 56 (K56) acetylation is an abundant modification of newly synthesized histone H3 molecules that are incorporated into chromosomes during S phase. Defects in the acetylation of K56 in histone H3 result in sensitivity to genotoxic agents that cause DNA strand breaks during replication. In the absence of DNA damage, the acetylation of histone H3 K56 largely disappears in G2. In contrast, cells with DNA breaks maintain high levels of acetylation, and the persistence of the modification is dependent on DNA damage checkpoint proteins. We suggest that the acetylation of histone H3 K56 creates a favourable chromatin environment for DNA repair and that a key component of the DNA damage response is to preserve this acetylation.

Examining minimal important change of the Canadian Occupational Performance Measure for subacute rehabilitation hospital inpatients.

BACKGROUND: The Canadian Occupational Performance Measure (COPM) is an individualized patient-reported outcome designed to evaluate the self-perceptions of a patient's occupational performance. Our study aimed to examine the minimal important change (MIC) in inpatients undergoing subacute rehabilitation. The MIC values were calculated using the three different anchor-based analyses with the transition index as an external criterion; the mean change method (MICMeanChange), the receiver operating characteristic (MICROC) analysis, and the predictive modeling method adjusted for the proportion of improved patients (MICadjust). In this study, the MICadjust value was considered as the most valid statistical method. We recruited 100 inpatients with various health conditions from subacute rehabilitation hospitals. Data were collected twice: an initial assessment and a reassessment one month later. The systematic interview format (Five Ws and How) was used for both the initial and second assessments to prevent information bias (response shift). RESULTS: Three patients who indicated deterioration on the transition index were excluded from all analyses, and 97 patients were analyzed in this study. The MICadjust values were 2.20 points (95% confidence interval 1.80-2.59) for the COPM performance score and 2.06 points (95% confidence interval 1.73-2.39) for the COPM satisfaction score. The MICMeanChange and MICROC values were considered less reasonable to interpret because the proportions of the improved patients subgroup were more than 50% (82.5%). CONCLUSIONS: The MICadjust value estimates from this study can help detect whether the patients' perceived occupational performance improved or did not change. The results support the multidisciplinary use of COPM in clinical practice and research on subacute rehabilitation inpatients.

Effect of home-based rehabilitation of purposeful activity-based electrical stimulation therapy for chronic stroke survivors: a crossover randomized controlled trial.

BACKGROUND: In this trial we combined the effect of purposeful activity and electrical stimulation therapy (PA-EST) to promote transition of severely hemiparetic upper limb to auxiliary upper limb in chronic stroke survivors in a single-case study. OBJECTIVE: The purpose of this study was to examine the effect of PA-EST on the upper limb motor function in a crossover randomized controlled trial. METHODS: The study included eight stroke survivors (age: 63.1±10.9 years) who were receiving home-based visiting occupational therapy. The average time since stroke onset was 8.8±5.6 years. All participants had severely hemiparetic upper limb, with the Fugl-Meyer Assessment upper extremity (FMA-U) score of 21.3±8.5. Participants were randomly assigned to group A or B. Group A received PA-EST for 3 months (phase 1), followed by standard stretching and exercise for 3 months (phase 2), whereas group B had the inverse order of treatments. To avoid carry-over effect, 1-month washout period was provided between the phase 1 and 2. Two-way analysis of variance (ANOVA) with repeated measures was used for the analysis. The primary outcome was FMA-U, and the secondary outcomes were, Motor Activity Log (MAL; amount of use [AOU] and quality of movement [QOM]), and Goal attainment scale-light (GAS-light). RESULTS: Repeated measures-ANOVA revealed a significant interaction between type of intervention and time for FMA-U (F = 16.303, P = 0.005), MAL AOU (F = 7.966, P = 0.026) and QOM (F = 6.408, P = 0.039), and GAS-light (F = 6.905, P = 0.034), where PA-EST was associated with significantly improved motor function and goal achievement compared with standard stretching. CONCLUSIONS: The PA-EST may have greater effects than stretch/exercise in the recovery of hand function as reflected in FMA-U, MAL, and GAS-light. Our results suggest that PA-EST is an important and useful home-based rehabilitation program for promoting the use of the severely hemiparetic upper limb in chronic stroke survivors.

Evaluation of changes in serum protein profiles during neoadjuvant chemotherapy in HER2-positive breast cancer using an LC-MALDI-TOF/MS procedure.

Comparison of protein profiles of sera acquired before and after preoperative chemotherapy for breast cancer may reveal tumor markers that could be used to monitor tumor response. In this study, we analyzed pre- and post-chemotherapy protein profiles of sera from 39 HER2-postive breast cancer patients (n=78 samples) who received 6 months of preoperative chemotherapy using LC-MALDI-TOF/MS technology. We detected qualitative and quantitative differences in pair-wise comparison of pre- and post chemotherapy samples that were different in patients who achieved pathological complete response (pCR, n=21) compared with those with residual disease (n=18). We identified 2329 and 3152 peaks as differentially expressed in the pre-chemotherapy samples of the responders and non-responders. Comparison of matching pre- and post-chemotherapy samples identified 34 (32 decreased, two increased) and 304 peaks (157 decreased, 147 increased) that significantly changed (p<0.01, false discovery rate ≤ 20%) after treatment in responders and non-responders, respectively. The top 11 most significantly altered peptide peaks with the greatest change in intensity were positively identified. These corresponded to eight proteins including α-2-macroglobulin, complement 3, hemopexin, and serum amyloid P in the responder group and chains C and A of apolipoprotein A-I, hemopexin precursor, complement C, and amyloid P component in the non-responding groups. All proteins decreased after therapy, except chain C apolipoprotein A and hemopexin precursor that increased. These results suggest that changes in serum protein levels occur in response to chemotherapy and these changes partly appear different in patients who are highly sensitive to chemotherapy compared with those with lesser response.

Development of a Japanese Version of a Collaborative Relationship Scale between Clients and Occupational Therapists.

OBJECTIVE: The success of a client-centred practice depends on the relationship between the client and therapist and on their ability to make constructive decisions together, particularly in the field of occupational therapy. The aim of this study was to develop a Occupational Therapy Collaborative Relationship Scale (OTCRS) to measure the quality of such interaction. METHODS: This work included constructing a draft questionnaire and testing its validity and reliability. A Rasch analysis was applied to determine its validity, and several tests were used to confirm its internal consistency. RESULTS: After reviewing more than 130 scientific papers and books, we built explicit selection criteria for issues to be addressed in this instrument, and we developed 40 questions to be included. These were analysed using a standard content validation process and a Rasch analysis to examine confirmation validity. A nine-item scale was finalised for testing (OTCRS-9). This review process revealed the validity, high internal consistency, and item/person separation reliability of OTCRS-9. CONCLUSION: This study presents only the initial phase of scale development. As suggested by the Consensus-based Standards for the Selection of Health Measurement Instruments (COSMIN), the OTCRS-9 score should be tested further for validity and reliability and should also be conducted in subjects of other ethnicities to improve its generalizability.

14-3-3 zeta protein secreted by tumor associated monocytes/macrophages from ascites of epithelial ovarian cancer patients.

Tumor associated monocytes/macrophages (MO/MA) are known contributors to the immune-inflammatory cell environment of advanced epithelial ovarian carcinoma (EOC). The secreted proteome of ascitic MO/MA was examined as an aid to the discovery of novel proteins in EOC that are likely to have biological relevance in the inflammatory pathways of EOC. Ascitic fluid MO/MA were isolated from EOC patients, grown short-term in serum-free media. MO/MA supernatants were analyzed for secreted proteins by HPLC fractionation followed by LC-tandem mass spectrometric analysis. The 14-3-3 zeta adaptor protein was identified in supernatants of three of three EOC patients but not in supernatants of buffy coat monocytes isolated from normal donors or the established monocyte cell line THP1. Moreover, 14-3-3 zeta was identified in ascitic fluids in eight of eight chemotherapy-naïve patients by both immunoblot and mass spectrometric analysis. Immunofluorescent staining for 14-3-3 zeta demonstrated expression of the protein on ascitic and peritumoral macrophages in EOC patients. 14-3-3 zeta was also expressed on endothelial cells in the peritumoral stroma and partially on tumor cells. Uptake of 14-3-3 zeta was observed in EOC cell lines co-cultured with the recombinant protein expressed in E. coli. It is demonstrated for the first time that the important adaptor protein 14-3-3 zeta is common to the secretome of ascitic MO/MA and the ascites of advanced EOC patients.

Stimulation of lung innate immunity protects against lethal pneumococcal pneumonia in mice.

RATIONALE: The lungs are a common site of serious infection in both healthy and immunocompromised subjects, and the most likely route of delivery of a bioterror agent. Since the airway epithelium shows great structural plasticity in response to inflammatory stimuli, we hypothesized it might also show functional plasticity. OBJECTIVES: To test the inducibility of lung defenses against bacterial challenge. METHODS: Mice were treated with an aerosolized lysate of ultraviolet-killed nontypeable (unencapsulated) Haemophilus influenzae (NTHi), then challenged with a lethal dose of live Streptococcus pneumoniae (Spn) delivered by aerosol. MEASUREMENTS AND MAIN RESULTS: Treatment with the NTHi lysate induced complete protection against challenge with a lethal dose of Spn if treatment preceded challenge by 4 to 24 hours. Lesser levels of protection occurred at shorter (83% at 2 h) and longer (83% at 48-72 h) intervals between treatment and challenge. There was also some protection when treatment was given 2 hours after challenge (survival increased from 14 to 57%), but not 24 hours after challenge. Protection did not depend on recruited neutrophils or resident mast cells and alveolar macrophages. Protection was specific to the airway route of infection, correlated in magnitude and time with rapid bacterial killing within the lungs, and was associated with increases of multiple antimicrobial polypeptides in lung lining fluid. CONCLUSIONS: We infer that protection derives from stimulation of local innate immune mechanisms, and that activated lung epithelium is the most likely cellular effector of this response. Augmentation of innate antimicrobial defenses of the lungs might have therapeutic value.

Proteomics-based identification of proteins secreted in apical surface fluid of squamous metaplastic human tracheobronchial epithelial cells cultured by three-dimensional organotypic air-liquid interface method.

Squamous cell carcinoma in the lung originates from bronchial epithelial cells that acquire increasingly abnormal phenotypes. Currently, no known biomarkers are clinically efficient for the early detection of premalignant lesions and lung cancer. We sought to identify secreted molecules produced from squamous bronchial epithelial cells cultured with organotypic culture methods. We analyzed protein expression patterns in the apical surface fluid (ASF) from aberrantly differentiated squamous metaplastic normal human tracheobronchial epithelial (NHTBE) and mucous NHTBE cells. Comparative two-dimensional PAGE analysis revealed 174 unique proteins in the ASF of squamous NHTBE cells compared with normal mucociliary differentiated NHTBE cells. Among them, 64 well-separated protein spots were identified by liquid chromatography-tandem mass spectrometry, revealing 22 different proteins in the ASF from squamous NHTBE cells. Expression of six of these proteins [SCC antigen 1 (SCCA1), SCC antigen 2 (SCCA2), S100A8, S100A9, Annexin I, and Annexin II] in the squamous NHTBE cells was further confirmed with immunoblot analysis. Notably, SCCA1 and SCCA2 were verified as being expressed in squamous metaplastic NHTBE cells but not in normal mucous NHTBE or normal bronchial epithelium. Moreover, SCCA1 and SCCA2 expression increased in in vitro lung carcinogenesis model cell lines with increasing malignancy. In summary, we identified proteins that are uniquely secreted from squamous metaplastic primary human bronchial epithelial cells cultured by the organotypic air-liquid interface method. These ASF proteins may be used to detect abnormal lesions in the lung without collecting invasive biopsy specimens.