Efficacy and safety of omalizumab in adult patients with wheat-dependent exercise-induced anaphylaxis: Reduction of in vitro basophil activation and allergic reaction to wheat.

BACKGROUND: In patients with wheat-dependent exercise-induced anaphylaxis (WDEIA), anaphylactic shock occurs frequently, therefore avoidance of wheat products is recommended. We aimed to evaluate efficacy and safety of long-term omalizumab treatment for adult patients with WDEIA. METHODS: In this phase 2, multicentre single-arm trial, 20 adult patients with WDEIA were enrolled (UMIN 000019250). All patients were administered 150-600 mg of omalizumab subcutaneously and evaluations (basophil activation and blood examination) were performed at regular intervals during administration period (0-48 weeks) and observation period (48-68 weeks). Primary endpoint was proportion of the patients who achieved a basophil activation rate below 10% with fractionated wheat preparations, and secondary endpoint was proportion of the patients with no allergic reactions after wheat products ingestion. RESULTS: During the omalizumab treatment, more than 80% of the patients achieved the basophil activation rate less than 10% against all fractionated wheat preparations, and 68.8% of the patients who achieved the primary endpoint experienced no allergic reaction. During the observation period, the proportion of the patients who achieved a basophil activation rate below 10% decreased gradually, and the proportion of patients with positive allergic reactions increased gradually thereafter and reached maximum of 46.7%. Severe adverse events were not observed during the study. CONCLUSIONS: Long-term omalizumab treatment is safe and effective for adult patients with WDEIA when assessed by basophil activation rate with wheat allergens as well as allergic reactions after lifting of restrictions on wheat intake. However, this is not enough to achieve desensitization.

High lymphocyte population-related predictive factors for a long-term response in non-small cell lung cancer patients treated with pemetrexed: a retrospective observational study.

BACKGROUND: Regimens combining pemetrexed (PEM) and immune checkpoint inhibitors (ICIs) targeting programmed cell death-1 (PD-1) or programmed death-ligand 1 (PD-L1) are widely used for the treatment of advanced non-squamous non-small-cell lung cancer (NSq-NSCLC). Recently, PEM was shown to induce immunogenic cell death (ICD) and to enhance immune-regulatory genes. Some patients demonstrate an extremely long-term response to PEM. It is possible that the continued response in these patients is dependent on not only the pharmacological induction of cytotoxic cell death but also antitumor immunity. However, factors that can predict outcomes associated with long-term PEM administration using blood test results have not yet been elucidated. We investigated the clinical characteristics and predictive factors in patients with advanced NSq-NSCLC who underwent long-term PEM maintenance therapy. METHODS: In total, 504 patients with advanced NSq-NSCLC who received PEM combination therapy/monotherapy (n = 414) or paclitaxel (PTX) combination therapy (n = 90) between January 2010 and November 2019 were recruited; 381 patients were retained for the final analysis. Patients treated with PEM (n = 301) were divided into subgroups according to the total cycles of PEM (≥ 17 [n = 25] for the long-term administration group and ≤ 16 [n = 276] for the intermediate/short-term group) and compared with another population (n = 80) treated with PTX combination regimen. We investigated clinical features and predictive biomarkers, focusing on immune-regulatory factors, absolute lymphocyte count (ALC), neutrophil-to-lymphocyte ratio (NLR), and PD-1 and PD-L1 expression, to predict long-term response to PEM. RESULTS: The long-term PEM administration group exhibited a higher ALC and a lower NLR than the shorter-term group did. Both these markers displayed greater association with progression-free survival and overall survival in the PEM combination therapy group than in the PTX combination therapy group. Increased PD-1 lymphocytes were associated with the long-term PEM response group, as PD-L1 expression in tumors was associated with a high incidence of immune-related adverse effects following ICI administration. CONCLUSIONS: ALC, NLR, and PD-1 expression are PEM-mediated predictive biomarkers that are indirectly related to tumor immunity and can provide useful predictive information on the long-term response to PEM in patients with NSq-NSCLC.

The synergetic effect of Imipenem-clarithromycin combination in the Mycobacteroides abscessus complex.

BACKGROUND: Nontuberculous mycobacteria (NTM) are ubiquitous organisms and the incidence of NTM infections has been increasing in recent years. Mycobacteroides abscessus (M. abscessus) is one of the most antimicrobial-resistant NTM; however, no reliable antibiotic regimen can be officially advocated. We evaluated the efficacy of clarithromycin in combination with various antimicrobial agents against the M. abscessus complex. RESULTS: Twenty-nine clinical strains of M. abscessus were isolated from various clinical samples. Of the isolates, 10 (34.5%) were of M. abscessus subsp. abscessus, 18 (62.1%) of M. abscessus subsp. massiliense, and 1 (3.4%) of M. abscessus subsp. bolletii. MICs of three antimicrobial agents (amikacin, imipenem, and moxifloxacin) were measured with or without clarithromycin. The imipenem-clarithromycin combination significantly reduced MICs compared to clarithromycin and imipenem monotherapies, including against resistant strains. The association between susceptibility of the M. abscessus complex and each combination of agents was significant (p = 0.001). Adjusted residuals indicated that the imipenem-clarithromycin combination had the synergistic effect (adjusted residual = 3.1) and suppressed the antagonistic effect (adjusted residual = - 3.1). In subspecies of M. abscessus complex, the association with susceptibility of M. abscessus subsp. massiliense was similarly statistically significant (p = 0.036: adjusted residuals of synergistic and antagonistic effect respectively: 2.6 and - 2.6). The association with susceptibility of M. abscessus subsp. abscessus also showed a similar trend but did not reach statistical significance. CONCLUSION: Our data suggest that the imipenem-clarithromycin combination could be the recommended therapeutic choice for the treatment of M. abscessus complex owing to its ability to restore antimicrobial susceptibility.

HLA-DQ and RBFOX1 as susceptibility genes for an outbreak of hydrolyzed wheat allergy.

BACKGROUND: Food allergy is a growing health problem worldwide because of its increasing prevalence, life-threatening potential, and shortage of effective preventive treatments. In an outbreak of wheat allergy in Japan, thousands of patients had allergic reactions to wheat after using soap containing hydrolyzed wheat protein (HWP). OBJECTIVES: The aim of the present study was to investigate genetic variation that can contribute to susceptibility to HWP allergy. METHODS: We conducted a genome-wide association study of HWP allergy in 452 cases and 2700 control subjects using 6.6 million genotyped or imputed single nucleotide polymorphisms. Replication was assessed by genotyping single nucleotide polymorphisms in independent samples comprising 45 patients with HWP allergy and 326 control subjects. RESULTS: Through the genome-wide association study, we identified significant associations with the class II HLA region on 6p21 (P = 2.16 × 10-24 for rs9271588 and P = 2.96 × 10-24 for HLA-DQα1 amino acid position 34) and with the RBFOX1 locus at 16p13 (rs74575857, P = 8.4 × 10-9). The associations were also confirmed in the replication data set. Both amino acid polymorphisms (HLA-DQß1 amino acid positions 13 and 26) located in the P4 binding pockets on the HLA-DQ molecule achieved the genome-wide significance level (P < 5.0 × 10-8). CONCLUSIONS: Our data provide the first demonstration of genetic risk for HWP allergy and show that this genetic risk is mainly represented by multiple combinations of HLA variants.

Specific Features of Fibrotic Lung Fibroblasts Highly Sensitive to Fibrotic Processes Mediated via TGF-ß-ERK5 Interaction.

BACKGROUND/AIMS: Lung fibrosis is associated with lung tissue contraction due to abnormal accumulation of myofibroblasts, which aggressively promote the fibrotic process. Transforming growth factor (TGF)-ß signaling in fibroblasts promotes extracellular matrix (ECM) synthesis and fibroblast migration and differentiation into myofibroblasts. Inhibition of extracellular signal-regulated kinase (ERK)5 blocks lung fibroblast activation by suppressing TGF-ß signaling. Here, we examined the effects of an ERK5 inhibitor on TGF-ß1-induced fibrosis in lung fibroblasts. METHODS: The effects of ERK5 inhibition following TGF-ß1 exposure were evaluated in lung fibroblasts isolated from fibrotic human lung tissues. Fibroblast-mediated collagen gel contraction and fibroblast migration towards fibronectin were assessed. Phenotypic differences in fibrotic fibroblasts were examined using the cap analysis gene expression method for genome-wide quantification of promoter activity. RESULTS: TGF-ß1stimulated contraction of collagen gels, fibroblast migration, and α-smooth muscle actin and fibronectin expression, and Smad3 phosphorylation were increased in fibrotic fibroblasts as compared to normal lung fibroblasts. Treatment with the ERK5 inhibitor blocked these responses to a greater extent in fibroblasts from patients with usual interstitial pneumonia as compared to nonspecific interstitial pneumonia, independent of bone morphogenetic protein/Smad1 regulation. Moreover, 223 genes including fibulin-5 -which is involved in the TGF-ß1-ERK5 signaling network- were upregulated in fibrotic fibroblasts, and ECM regulation was found to be enriched in the Reactome analysis. CONCLUSION: ERK5 inhibition attenuated the high sensitivity of fibrotic fibroblasts to TGF-ß1/Smad3 signaling. Thus, the ERK5 pathway components and fibulin-5 are potential therapeutic targets to prevent lung fibrosis progression.

Pirfenidone attenuates lung fibrotic fibroblast responses to transforming growth factor-ß1.

BACKGROUND: Pirfenidone, an antifibrotic agent used for the treatment of idiopathic pulmonary fibrosis (IPF), functions by inhibiting myofibroblast differentiation, which is involved in transforming growth factor (TGF)-ß1-induced IPF pathogenesis. However, unlike normal lung fibroblasts, the relationship between pirfenidone responses of TGF-ß1-induced human fibrotic lung fibroblasts and lung fibrosis has not been elucidated. METHODS: The effects of pirfenidone were evaluated in lung fibroblasts isolated from fibrotic human lung tissues after TGF-ß1 exposure. The ability of two new pharmacological targets of pirfenidone, collagen triple helix repeat containing protein 1(CTHRC1) and four-and-a-half LIM domain protein 2 (FHL2), to mediate contraction of collagen gels and migration toward fibronectin were assessed in vitro. RESULTS: Compared to control lung fibroblasts, pirfenidone significantly restored TGF-ß1-stimulated fibroblast-mediated collagen gel contraction, migration, and CTHRC1 release in lung fibrotic fibroblasts. Furthermore, pirfenidone attenuated TGF-ß1- and CTHRC1-induced fibroblast activity, upregulation of bone morphogenic protein-4(BMP-4)/Gremlin1, and downregulation of α-smooth muscle actin, fibronectin, and FHL2, similar to that observed post-CTHRC1 inhibition. In contrast, FHL2 inhibition suppressed migration and fibronectin expression, but did not downregulate CTHRC1. CONCLUSIONS: Overall, pirfenidone suppressed fibrotic fibroblast-mediated fibrotic processes via inverse regulation of CTHRC1-induced lung fibroblast activity. Thus, CTHRC1 can be used for predicting pirfenidone response and developing new therapeutic targets for lung fibrosis.

A novel technique to diagnose non-melanoma skin cancer by thermal conductivity measurements: Correlations with cancer stromal factors.

The skin surface temperature reflects the physiological state of the human body. Quantitative methods of identification of skin cancers based on accurate measurement of effective thermal conductivity (ETC) are among the promising diagnostic tools for differentiating non-invasive and invasive melanomas before surgical treatment. To validate these findings, in this report, the diagnostic methods for invasive and non-invasive extramammary Paget's disease (EMPD) and squamous cell carcinoma (SCC) were further tested by measuring the absolute value of skin surface temperature and the ETC of the skin. In addition, to investigate the stromal factors that might affect ETC, immunohistochemical staining for LL37, periostin (POSTN), MMP12, and MMP28 was performed. The invasive SCC and EMPD group showed a relatively higher skin surface temperature compared to the in situ SCC group. The non-invasive EMPD and SCC group showed significantly lower values of ETC at lesions, whereas the invasive EMPD group showed significantly higher ETC values at lesions compared to healthy skin. Immunohistochemical staining showed that the percentage of LL37-producing cells was significantly increased in invasive EMPD and SCC compared to that in non-invasive EMPD and SCC. Moreover, Spearman's rank correlation test showed a significant inverse correlation between the percentage of MMP12-positive cells and increased levels of ETC-expressing areas in EMPD and SCC (r = -.5997). The present study suggested that differences in ETC could be a novel high-accuracy diagnostic technique for non-melanoma skin cancer, especially for detecting dermal invasion of SCC and EMPD.

Delineation of solution burst-phase protein folding events by encapsulating the proteins in silica gels.

Many studies have shown that during the early stages of the folding of a protein, chain collapse and secondary structure formation lead to a partially folded intermediate. Thus, direct observation of these early folding events is crucial if we are to understand protein-folding mechanisms. Notably, these events usually manifest as the initial unresolvable signals, denoted the burst phase, when monitored during conventional mixing experiments. However, folding events can be substantially slowed by first trapping a protein within a silica gel with a large water content, in which the trapped native state retains its solution conformation. In this study, we monitored the early folding events involving secondary structure formation of five globular proteins, horse heart cytochrome c, equine ß-lactoglobulin, human tear lipocalin, bovine α-lactalbumin, and hen egg lysozyme, in silica gels containing 80% (w/w) water by CD spectroscopy. The folding rates decreased for each of the proteins, which allowed for direct observation of the initial folding transitions, equivalent to the solution burst phase. The formation of each initial intermediate state exhibited single exponential kinetics and Arrhenius activation energies of 14-31 kJ/mol.

Relationship between chain collapse and secondary structure formation in a partially folded protein.

Chain collapse and secondary structure formation are frequently observed during the early stages of protein folding. Is the chain collapse brought about by interactions between secondary structure units or is it due to polymer behavior in a poor solvent (coil-globule transition)? To answer this question, we measured small-angle X-ray scattering for a series of ß-lactoglobulin mutants under conditions in which they assume a partially folded state analogous to the folding intermediates. Mutants that were designed to disrupt the secondary structure units showed the gyration radii similar to that of the wild type protein, indicating that chain collapse is due to coil-globule transitions.

Novel conditionally replicating adenovirus-mediated efficient detection of circulating tumor cells in lung cancer patients.

Circulating tumor cells (CTCs) are present in the blood of cancer patients from the early stage of cancer development, and their presence has been correlated with patient prognosis and treatment responses. Accordingly, CTCs have been attracting attention as a novel biomarker for early detection of cancer and monitoring of treatment responses. However, since patients typically have only a few CTCs per milliliter of blood, development of an accurate and highly sensitive CTC detection method is crucial. We previously developed a CTC detection method using a novel conditionally replicating adenovirus (Ad) that expresses green fluorescence protein (GFP) in a tumor cell-specific manner by expressing the E1 gene using a tumor-specific human telomerase reverse transcriptase (hTERT) promoter (rAdF35-142T-GFP). CTCs were efficiently detected using rAdF35-142T-GFP, but GFP expression levels in the CTCs and production efficiencies of rAdF35-142T-GFP were relatively low. In this study, in order to overcome these problems, we developed four types of novel GFP-expressing conditionally replicating Ads and examined their ability to visualize CTCs in the blood samples of lung cancer patients. Among the four types of novel recombinant Ads, the novel conditionally replicating Ad containing the 2A peptide and the GFP gene downstream of the E1A gene and the adenovirus death protein (ADP) gene in the E3 region (rAdF35-E1-2A-GFP-ADP) mediated the highest number of GFP-positive cells in the human cultured tumor cell lines. Titers of rAdF35-E1-2A-GFP-ADP were significantly higher (about 4-fold) than those of rAdF35-142T-GFP. rAdF35-E1-2A-GFP-ADP and rAdF35-142T-GFP efficiently detected CTCs in the blood of lung cancer patients at similar levels. GFP+/CD45- cells (CTCs) were found in 10 of 17 patients (58.8%) for both types of recombinant Ads.

The synergetic effect of sitafloxacin-arbekacin combination in the Mycobacterium abscessus species.

Mycobacterium abscessus species (MABS) is the most commonly isolated rapidly growing mycobacteria (RGM) and is one of the most antibiotic-resistant RGM with rapid progression, therefore, treatment of MABS is still challenging. We here presented a new combination treatment with sitafloxacin that targeted rough morphotypes of MABS, causing aggressive infections. Thirty-four clinical strains of MABS were isolated from various clinical samples at the Juntendo university hospital from 2011 to 2020. The susceptibility to a combination of sitafloxacin and antimicrobial agents was compared to that of the antimicrobial agents alone. Out of 34 MABS, 8 strains treated with sitafloxacin-amikacin combination, 9 of sitafloxacin-imipenem combination, 19 of sitafloxacin-arbekacin combination, and 9 of sitafloxacin-clarithromycin combination showed synergistic effects, respectively. Sitafloxacin-arbekacin combination also exhibited the synergistic effects against 10 of 22 Mycobacterium abscessus subspecies massiliense (Mma) strains and 8 of 11 Mycobacterium abscessus subspecies abscessus (Mab) strains, a highly resistant subspecies of MABS. The sitafloxacin-arbekacin combination revealed more synergistic effects in rough morphotypes of MABS (p = 0.008). We demonstrated the synergistic effect of the sitafloxacin-arbekacin combination against MABS. Further, this combination regimen might be more effective against Mab or rough morphotypes of MABS.

Endogenous retinoids in mammalian growth plate cartilage: analysis and roles in matrix homeostasis and turnover.

The growth plate contains resting and proliferating chondrocytes in its upper zones (UGP) and maturing and hypertrophic chondrocytes in its lower zones (LGP), but the mechanisms by which it operates to sustain skeletal growth are not fully clear. Retinoid signaling was previously found to be nearly absent in UGP, but to be much stronger in LGP coincident with hypertrophy, extracellular matrix turnover and endochondral bone formation. To determine whether such distinct signaling levels and phenotypic events reflect different endogenous retinoid levels, the upper two-thirds and lower one-third of rabbit rib growth plates were microsurgically isolated and processed for ultrasensitive retinoid LC-tandem MS quantification. Indeed, the UGP samples contained only about a 0.6 nm concentration of all-trans-retinoic acid (atRA) that is the most active natural retinoid in tissues, whereas LGP samples contained nearly 3-fold higher atRA levels (about 1.8 nM). Perichondrium was quite rich in atRA (about 4.9 nM). Interestingly, the levels of retinol, the major but inactive atRA precursor, were similar in all tissues (1.1-1.6 µM), suggesting that the distinct atRA levels in UGP and LGP reflect different retinoid anabolic capacity. Indeed, RALDH2 and CRABP1 transcript levels were much higher in LGP than UGP samples. To determine the minimum effective atRA concentration, chondrogenic cells transfected with a retinoic acid response element (RARE)-luc reporter plasmid were treated with different concentrations of exogenous atRA (0-100 nM). About 3 nm atRA was needed to elicit appreciable RARE-luc reporter activity and to decrease proteoglycan synthesis and activity of an aggrecan enhancer reporter plasmid. In sum, the data indicate that (i) the endogenous levels of atRA are significantly higher in hypertrophic than upper zones of growth plate; (ii) such difference likely reflects distinct retinoid anabolic capacity; and (iii) importantly, atRA levels in hypertrophic portion are within effective ranges to elicit retinoid signaling and action, but those in upper zones are not.

Construction of osteochondral-like tissue graft combining ß-tricalcium phosphate block and scaffold-free centrifuged chondrocyte cell sheet.

BACKGROUND: The combination of a ß-tricalcium phosphate (ßTCP) block with a scaffold-free chondrocyte sheet formed by the centrifugation of chondrocytes in a well was investigated with the aim of constructing an osteochondral-like structure. METHODS: Human and porcine articular cartilage chondrocytes were respectively centrifuged in a 96-well plate or cell culture insert (0.32 cm(2)) that was set in a 24-well plate, cultivated in the respective vessel for 3 weeks, and the cell sheets were harvested. In some cases, a cylindrical ßTCP block (diameter 5 mm, height 3 mm) was placed on the sheet on days 1-7. The sheet size, cell number, and sulfated glycosaminoglycan accumulation were determined. RESULTS: The use of a 96-well plate for not suspension but adhesion culture and the initial centrifugation of a well containing cells were crucial to obtaining a uniformly thick cell sheet. The glycosaminoglycan density of the harvested cell sheet was comparable to that of the pellet culture. An inoculum cell number of more than 31 × 10(5) cells tended to result in a curved cell sheet. Culture involving 18.6 × 10(5) cells and the 96-well plate for adhesion culture showed no curving of the cell sheet (thickness of 0.85 mm), and these were found to be the best of the culture conditions tested. The timing of the addition of a ßTCP block to the cell sheet (1-7 days) markedly affected the balance between the thickness of cell sheet parts on and in the ßTCP block. CONCLUSION: Centrifugation and subsequent cultivation of chondrocytes (18.6 × 10(5) cells) in a 96-well plate for adhesion culture led to the production of a scaffold-free cartilage-like cell sheet with a thickness of 0.85 mm. A combined osteochondral-like structure was produced by putting a ßTCP block on the cell sheet. The thickness of the cell sheet on the ßTCP block and the binding strength between the cell sheet and the ßTCP block could be optimized by adjusting the inoculum cell number and timing of ßTCP block addition.

Cancer-associated fibroblast migration in non-small cell lung cancers is modulated by increased integrin α11 expression.

Cancer-associated fibroblasts (CAFs) regulate cancer progression through the modulation of extracellular matrix (ECM) and cancer cell adhesion. While undergoing a series of phenotypic changes, CAFs control cancer-stroma interactions through integrin receptor signaling. Here, we isolated CAFs from patients with non-small-cell lung cancer (NSCLC) and examined their gene expression profiles. We identified collagen type XI α1 (COL11A1), integrin α11 (ITGA11), and the ITGA11 major ligand collagen type I α1 (COL1A1) among the 390 genes that were significantly enriched in NSCLC-associated CAFs. Increased ITGA11 expression in cancer stroma was correlated with a poor clinical outcome in patients with NSCLC. Increased expression of fibronectin and collagen type I induced ITGA11 expression in CAFs. The cellular migration of CAFs toward collagen type I and fibronectin was promoted via ERK1/2 signaling, independently of the fibronectin receptor integrin α5ß1. Additionally, ERK1/2 signaling induced ITGA11 and COL11A1 expression in cancer stroma. We, therefore, propose that targeting ITGA11 and COL11A1 expressing CAFs to block cancer-stroma interactions may serve as a novel, promising anti-tumor strategy.

Retinoic acid receptors are required for skeletal growth, matrix homeostasis and growth plate function in postnatal mouse.

The retinoic acid receptors alpha, beta and gamma (RARalpha, RARbeta and RARgamma) are nuclear hormone receptors that regulate fundamental processes during embryogenesis, but their roles in skeletal development and growth remain unclear. To study skeletal-specific RAR function, we created conditional mouse mutants deficient in RAR expression in cartilage. We find that mice deficient in RARalpha and RARgamma (or RARbeta and RARgamma) exhibit severe growth retardation obvious by about 3 weeks postnatally. Their growth plates are defective and, importantly, display a major drop in aggrecan expression and content. Mice deficient in RARalpha and RARbeta, however, are virtually normal, suggesting that RARgamma is essential. In good correlation, we find that RARgamma is the most strongly expressed RAR in mouse growth plate and its expression characterizes the proliferative and pre-hypertrophic zones where aggrecan is strongly expressed also. By being avascular, those zones lack endogenous retinoids as indicated by previous RARE reporter mice and our direct biochemical measurements and thus, RARgamma is likely to exert ligand-less repressor function. Indeed, our data indicate that: aggrecan production is enhanced by RARgamma over-expression in chondrocytes under retinoid-free culture conditions; production is further boosted by co-repressor Zac1 or pharmacologic agents that enhance RAR repressor function; and RAR/Zac1 function on aggrecan expression may involve Sox proteins. In sum, our data reveal that RARs, and RARgamma in particular, exert previously unappreciated roles in growth plate function and skeletal growth and regulate aggrecan expression and content. Since aggrecan is critical for growth plate function, its deficiency in RAR-mutant mice is likely to have contributed directly to their growth retardation.

Articular cartilage repair with autologous bone marrow mesenchymal cells.

Articular cartilage defects that do not repair spontaneously induce osteoarthritic changes in joints over a long period of observation. In this study, we examined the usefulness of transplanting culture-expanded bone marrow mesenchymal cells into osteochondral defects of joints with cartilage defects. First, we performed experiments on rabbits and up on obtaining good results proceeded to perform the experiments on humans. Macroscopic and histological repair with this method was good, and good clinical results were obtained although there was no significant difference with the control group. Recent reports have indicated that this procedure is comparable to autologous chondrocyte implantation, and concluded that it was a good procedure because it required one step less than that required by surgery, reduced costs for patients, and minimized donor site morbidity. Although some reports have previously shown that progenitor cells formed a tumor when implanted into immune-deficient mice after long term in vitro culture, the safety of the cell transplantation was confirmed by our clinical experience. Thus, this procedure is useful, effective, and safe, but the repaired tissues were not always hyaline cartilage. To obtain better repair with this procedure, treatment approaches using some growth factors during in vitro culture or gene transfection are being explored.

Highly sensitive ELISA for determining serum keratan sulphate levels in the diagnosis of OA.

OBJECTIVE: There have been a large number of reports on alterations in the serum level of keratan sulphate (KS), a potential marker of articular cartilage degeneration in patients with arthropathy. Such studies have commonly employed ELISA using the anti-KS monoclonal antibody 1/20/5D4 (5D4-ELISA) to determine KS levels. Recently, a highly sensitive KS ELISA (HS-ELISA) kit has been developed, allowing determination of serum KS levels even in small animals, which were formerly undetectable with 5D4-ELISA. However, the effectiveness of this kit in humans has not been demonstrated. The objective of this study was to assess the usefulness of the HS-ELISA for the analysis of human serum samples. METHODS: Serum samples were collected from 28 patients with knee OA and 23 healthy volunteers. KS was determined by 5D4-ELISA and HS-ELISA, and measurements were compared with those obtained by HPLC. KS levels in serum samples with protease pretreatment were also determined by HS-ELISA. RESULTS: KS levels determined by HS-ELISA exhibited a better correlation with those determined by HPLC, and a higher diagnostic sensitivity for OA compared with 5D4-ELISA. Protease pretreatment of serum further improved the correlation between the values obtained by HS-ELISA and HPLC, as well as the diagnostic sensitivity of HS-ELISA for OA. CONCLUSIONS: HS-ELISA proved useful for determining KS level in serum and the diagnosis of OA. Pretreatment of serum samples with a protease further improved the performance of HS-ELISA.

Hierarchical Gaussian Descriptors with Application to Person Re-Identification.

Describing the color and textural information of a person image is one of the most crucial aspects of person re-identification (re-id). Although a covariance descriptor has been successfully applied to person re-id, it loses the local structure of a region and mean information of pixel features, both of which tend to be the major discriminative information for person re-id. In this paper, we present novel meta-descriptors based on a hierarchical Gaussian distribution of pixel features, in which both mean and covariance information are included in patch and region level descriptions. More specifically, the region is modeled as a set of multiple Gaussian distributions, each of which represents the appearance of a local patch. The characteristics of the set of Gaussian distributions are again described by another Gaussian distribution. Because the space of Gaussian distribution is not a linear space, we embed the parameters of the distribution into a point of Symmetric Positive Definite (SPD) matrix manifold in both steps. We show, for the first time, that normalizing the scale of the SPD matrix enhances the hierarchical feature representation on this manifold. Additionally, we develop feature norm normalization methods with the ability to alleviate the biased trends that exist on the SPD matrix descriptors. The experimental results conducted on five public datasets indicate the effectiveness of the proposed descriptors and the two types of normalizations.

Numerical Simulation of Effects of Bioheat Transfer Characteristics of Malignant Melanoma on Thermal Conductivity Measurements.

In our previous study, we successfully detected a difference in the effective thermal conductivity between an invasive melanoma lesion and healthy skin, through clinical experiments conducted on melanoma patients. We found that the effective thermal conductivity of the lesions correlated with the tumor thickness, suggesting that it may be correlated with the prognostic risk of melanoma. However, the bioheat transfer mechanisms of the correlation remained unknown. The aim of this study was to numerically investigate the effects of the bioheat transfer characteristics of malignant melanoma on thermal conductivity measurements and explore the cause of the difference in the effective thermal conductivity between lesions and healthy skin. We used two different bioheat transfer models, the Pennes model and local thermal nonequilibrium model, and investigated the cause of the aforementioned differences by varying the bioheat transfer parameters associated with the thermophysical properties and blood flow of a tumor. The calculation results indicated that the contribution of the blood flow can be dominant in a measurement comprising the use of a guard-heated thermistor probe. Therefore, we found that it is necessary to take into consideration the contribution of the convective term to the effective thermal conductivity of the lesion in order to explain the clinical data of a Stage IV invasive melanoma.

Mesenchymal Characteristics and Predictive Biomarkers on Circulating Tumor Cells for Therapeutic Strategy.

Metastasis-related events are the primary cause of cancer-related deaths, and circulating tumor cells (CTCs) have a pivotal role in metastatic relapse. CTCs include a variety of subtypes with different functional characteristics. Interestingly, the epithelial-mesenchymal transition (EMT) markers expressed in CTCs are strongly associated with poor clinical outcome and related to the acquisition of circulating tumor stem cell (CTSC) features. Recent studies have revealed the existence of CTC clusters, also called circulating tumor microemboli (CTM), which have a high metastatic potential. In this review, we present current opinions regarding the clinical significance of CTCs and CTM with a mesenchymal phenotype as clinical surrogate markers, and we summarize the therapeutic strategy according to phenotype characterization of CTCs in various types of cancers for future precision medicine.