Maternal dendritic cells influence fetal allograft response following murine in-utero hematopoietic stem cell transplantation.

BACKGROUND: Intrauterine hematopoietic stem cell transplantation (IUT), potentially curative in congenital haematological disease, is often inhibited by deleterious immune responses to donor cells resulting in subtherapeutic donor cell chimerism (DCC). Microchimerism of maternal immune cells (MMc) trafficked into transplanted recipients across the placenta may directly influence donor-specific alloresponsiveness, limiting DCC. We hypothesized that dendritic cells (DC) among trafficked MMc influence the development of tolerogenic or immunogenic responses towards donor cells, and investigated if maternal DC-depletion reduced recipient alloresponsiveness and enhanced DCC. METHODS: Using transgenic CD11c.DTR (C57BL/6) female mice enabled transient maternal DC-depletion with a single dose of diphtheria toxin (DT). CD11c.DTR females and BALB/c males were cross-mated, producing hybrid pups. IUT was performed at E14 following maternal DT administration 24 h prior. Bone marrow-derived mononuclear cells were transplanted, obtained from semi-allogenic BALB/c (paternal-derived; pIUT), C57BL/6 (maternal-derived; mIUT), or fully allogenic (aIUT) C3H donor mice. Recipient F1 pups were analyzed for DCC, while maternal and IUT-recipient immune cell profile and reactivity were examined via mixed lymphocyte reactivity functional assays. T- and B-cell receptor repertoire diversity in maternal and recipient cells were examined following donor cell exposure. RESULTS: DCC was highest and MMc was lowest following pIUT. In contrast, aIUT recipients had the lowest DCC and the highest MMc. In groups that were not DC-depleted, maternal cells trafficked post-IUT displayed reduced TCR & BCR clonotype diversity, while clonotype diversity was restored when dams were DC-depleted. Additionally, recipients displayed increased expression of regulatory T-cells and immune-inhibitory proteins, with reduced proinflammatory cytokine and donor-specific antibody production. DC-depletion did not impact initial donor chimerism. Postnatal transplantation without immunosuppression of paternal donor cells did not increase DCC in pIUT recipients; however there were no donor-specific antibody production or immune cell changes. CONCLUSIONS: Though maternal DC depletion did not improve DCC, we show for the first time that MMc influences donor-specific alloresponsiveness, possibly by expanding alloreactive clonotypes, and depleting maternal DC promotes and maintains acquired tolerance to donor cells independent of DCC, presenting a novel approach to enhancing donor cell tolerance following IUT. This may have value when planning repeat HSC transplantations to treat haemoglobinopathies.

Rapid Detection of Pathogenic Bacteria by the Naked Eye.

Escherichia coli O157:H7 and Staphylococcus aureus are common pathogens. Gram-negative bacteria, such as E. coli, contain high concentrations of endogenous peroxidases, whereas Gram-positive bacteria, such as S. aureus, possess abundant endogenous catalases. Colorless 3,5,3',5'-tetramethyl benzidine (TMB) changes to blue oxidized TMB in the presence of E. coli and a low concentration of H2O2 (e.g., ~11 mM) at pH of 3. Moreover, visible air bubbles containing oxygen are generated after S. aureus reacts with H2O2 at a high concentration (e.g., 180 mM) at pH of 3. A novel method for rapidly detecting the presence of bacteria on the surfaces of samples, on the basis of these two endogenous enzymatic reactions, was explored. Briefly, a cotton swab was used for collecting bacteria from the surfaces of samples, such as tomatoes and door handles, then two-step endogenous enzymatic reactions were carried out. In the first step, a cotton swab containing bacteria was immersed in a reagent comprising H2O2 (11.2 mM) and TMB for 25 min. In the second step, the swab was dipped further in H2O2 (180 mM) at pH 3 for 5 min. Results showed that the presence of Gram-negative bacteria, such as E. coli with a cell number of ≥ ~105, and Gram-positive bacteria, such as S. aureus with a cell number of ≥ ~106, can be visually confirmed according to the appearance of the blue color in the swab and the formation of air bubbles in the reagent solution, respectively, within ~30 min. To improve visual sensitivity, we dipped the swab carrying the bacteria in a vial containing a growth broth, incubated it for ~4 h, and carried out the two-stage reaction steps. Results showed that bluish swabs resulting from the presence of E. coli O157: H7 with initial cell numbers of ≥ ~34 were obtained, whereas air bubbles were visible in the samples containing S. aureus with initial cell numbers of ≥ ~8.5 × 103.

Functional magnetic nanoparticle-based affinity probe for MALDI mass spectrometric detection of ricin B.

The use of lactosylated Fe3O4 magnetic nanoparticles (MNP@LAC) has been explored as affinity probes against ricin B based on galactose-ricin B binding interactions. Lactose was bound onto the surface of aminated MNPs through the Maillard reaction. The enrichment of ricin B took ~1 h by incubating MNP@LAC with samples under shaking at room temperature, followed by magnetic isolation. The resultant MNP@LAC-ricin B conjugates were characterized by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The limit of detection toward ricin B was ~3 nM by using the developed method. It was possible to detect the peptides derived from the tryptic digest of trace ricin B (~0.39 nM) enriched by the MNP@LAC probes followed by tryptic digestion and MALDI-MS analysis. The feasibility of using the developed method for detection of ricin B from complex white corn starch samples spiked with trace ricin B was demonstrated.

Magnetic Graphene Oxide-Based Affinity Surface-Assisted Laser Desorption/Ionization Mass Spectrometry for Screening of Aflatoxin B1 from Complex Samples.

Aflatoxin B1 (AFB1), commonly found in agriculture products, has been considered as a carcinogen. Thus, to develop analytical methods that can be used to rapidly screen the presence of AFB1 in complex samples is important. Surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) uses inorganic materials as assisting materials to facilitate desorption/ionization of analytes. The feasibility of using GO as the affinity probe against AFB1 and as the assisting material in SALDI-MS analysis was first demonstrated. We also explored a facile method to impose magnetism on GO to generate magnetic GO (MGO) nanoprobes by simply incubating GO in aqueous FeCl3 under microwave heating. The generated MGO nanoprobes possessed magnetism and were capable of enriching trace AFB1 from complex samples. AFB1 enrichment took only 6 min by incubating MGO with samples under microwave heating (power = 90 W). Followed by magnetic isolation, the isolated conjugates were ready for SALDI-MS analysis. The enrichment steps including trapping and isolation can be completed within ∼10 min. The lowest detectable concentration of our method toward AFB1 was ∼1 nM. Results also showed that AFB1 can be selectively detected from complex samples, including cell lysates of fungal spores, AFB1-spiked peanut, and wheat samples, by using the developed method. The selectivity of our method against AFB1 from the samples containing other toxins including aflatoxin G1 and ochratoxin A was also examined. According to these results, we believe that the developed method should have the potential to be used for rapid screening of AFB1 from real-world samples.

Maternal microchimerism and cell-mediated immune-modulation enhance engraftment following semi-allogenic intrauterine transplantation.

Successful intrauterine hematopoietic cell transplantation (IUT) for congenital hemoglobinopathies is hampered by maternal alloresponsiveness. We investigate these interactions in semi-allogenic murine IUT. E14 fetuses (B6 females × BALB/c males) were each treated with 5E+6 maternal (B6) or paternal (BALB/c) bone marrow cells and serially monitored for chimerism (>1% engraftment), trafficked maternal immune cells, and immune responsiveness to donor cells. A total of 41.0% of maternal IUT recipients (mIUT) were chimeras (mean donor chimerism 3.0 ± 1.3%) versus 75.0% of paternal IUT recipients (pIUT, 3.6 ± 1.1%). Chimeras showed higher maternal microchimerism of CD4, CD8, and CD19 than non-chimeras. These maternal cells showed minimal responsiveness to B6 or BALB/c stimulation. To interrogate tolerance, mIUT were injected postnatally with 5E+6 B6 cells/pup; pIUT received BALB/c cells. IUT-treated pups showed no changes in trafficked maternal or fetal immune cell levels compared to controls. Donor-specific IgM and IgG were expressed by 1%-3% of recipients. mIUT splenocytes showed greater proliferation of regulatory T cells (Treg) upon BALB/c stimulation, while B6 stimulation upregulated the pro-inflammatory cytokines more than BALB/c. pIUT splenocytes produced identical Treg and cytokine responses to BALB/c and B6 cells, with higher Treg activity and lower pro-inflammatory cytokine expression upon exposure to BALB/c. In contrast, naïve fetal splenocytes demonstrated greater alloresponsiveness to BALB/c compared to B6 cells. Thus pIUT, associated with increased maternal cell trafficking, modulates fetal Treg, and cytokine responsiveness to donor cells more efficiently than mIUT, resulting in improved engraftment. Paternal donor cells may be considered alternatively to maternal donor cells for intrauterine and postnatal transplantation to induce tolerance and maintain engraftment.

Using lactosylated cysteine functionalized gold nanoparticles as colorimetric sensing probes for rapid detection of the ricin B chain.

A new colorimetric method that can be used to rapidly detect toxic ricin is demonstrated. Lactosylated cysteine-functionalized gold nanoparticles (Au@LACY NPs) were prepared by a one-pot reaction and employed as optical probes for determination of ricin B chain. It is found that the Au@LACY NPs undergo aggregation in the presence of ricin B chain. This leads to surface plasmon coupling effects of the particles and a color change from red to blue, with absorption maxima at 519 and 670 nm, respectively. The feasibility of using the current approach for quantitative analysis of ricin B chain is also demonstrated. The calibration plot is generated by plotting the ratio of the absorbance at the wavelength of 634 to 518 nm versus the concentration of the ricin B chain. The spectrophotometric method has a ~29 pM (~ 0.91 ng·mL-1) detection limit, and the sample with the concentration of ~ 400 pM (~ 13 ng·mL-1) can be detected visually. Graphical abstractSchematic representation of using lactosylated cysteine capped gold nanoparticles (Au@LACY NPs) as colorimetric probes for the ricin B chain through surface plasmon coupling effects. Sample solution turns from red to blue in the presence of ricin B chain.

Thin peptide hydrogel membranes suitable as scaffolds for engineering layered biostructures.

A short tetramer peptide, Ac-IVKC, spontaneously formed a hydrogel in water. Disulfide bonds were introduced via hydrogen peroxide (H2O2)-assisted oxidation, resulting in (Ac-IVKC)2 dimers. The extent of disulfide bond formation and gel stiffness increased with the amount of H2O2 used and 100% dimerization was achieved with 0.2% H2O2. The resultant gel achieved an elastic modulus of ∼0.9 MPa, which to our knowledge, has not been reported for peptide-based hydrogels. The enhanced mechanical property enabled the fabrication of thin and transparent membranes. The hydrogel could also be handled with forceps at mm thickness, greatly increasing its ease of physical manipulation. Excess H2O2 was removed and the membrane was then infused with cell culture media. Various cells, including primary human corneal stromal and epithelial cells, were seeded onto the hydrogel membrane and demonstrated to remain viable. Depending on the intended application, specific cell combination or membrane stacking order could be used to engineer layered biostructures. STATEMENT OF SIGNIFICANCE: A short tetramer peptide - Ac-IVKC - spontaneously formed a hydrogel in water and disulfide bonds were introduced via hydrogen peroxide (H2O2)-assisted oxidation. The extent of disulfide-bond formation and gel stiffness were modulated by the amount of H2O2. At maximum disulfide-bond formation, the hydrogel achieved an elastic modulus of ∼0.9 MPa, which to our knowledge, has not been reported for peptide-based hydrogels. The enhanced mechanical property enabled the fabrication of thin transparent membranes that can be physically manipulated at mm thickness. The gels also supported 3D cell growth, including primary human corneal stromal and epithelial cells. Depending on the intended application, specific combination of cells or individual membrane stacking order could be used to engineer layered biostructures.

Ultrathin, Strong, and Cell-Adhesive Agarose-Based Membranes Engineered as Substrates for Corneal Endothelial Cells.

We aimed to bioengineer a scaffold that can facilitate the transplantation of corneal endothelial cells (CEC), given the global shortage of cadaveric donor tissues. Although agarose (A) has outstanding biocompatibility and mechanical properties, it natively does not permit cell adhesion. In this study, agarose was modified with different attachment signals: GRGD (giving AR as product), lysine (AK), poly lysine (AP), and fish-derived gelatin (AG). Samples with varying conjugation ratios were prepared. All products formed bulk hydrogels, which were then collapsed into ultrathin membranes in a controlled environment. Membranes were evaluated for their ability to support attachment of various cell types. Cells, however, preferred the AG series of membrane. Notably, primary rabbit CEC remained attached and viable for ⩾4 weeks. The cells also stained positive for CD166, ZO-1 and Na+/K+ ATPase, indicative of function. The hydrated AG membranes allowed >96% transmittance of visible light. The membranes were typically ∼15 µm thick and did not swell significantly after immersion in PBS. Tensile strength was 49-60 MPa, while young's modulus was 525-596 MPa. This membrane thus offers great promise as a scaffold for CEC during endothelial keratoplasty.

Prediction of drug-induced nephrotoxicity and injury mechanisms with human induced pluripotent stem cell-derived cells and machine learning methods.

The renal proximal tubule is a main target for drug-induced toxicity. The prediction of proximal tubular toxicity during drug development remains difficult. Any in vitro methods based on induced pluripotent stem cell-derived renal cells had not been developed, so far. Here, we developed a rapid 1-step protocol for the differentiation of human induced pluripotent stem cells (hiPSC) into proximal tubular-like cells. These proximal tubular-like cells had a purity of >90% after 8 days of differentiation and could be directly applied for compound screening. The nephrotoxicity prediction performance of the cells was determined by evaluating their responses to 30 compounds. The results were automatically determined using a machine learning algorithm called random forest. In this way, proximal tubular toxicity in humans could be predicted with 99.8% training accuracy and 87.0% test accuracy. Further, we studied the underlying mechanisms of injury and drug-induced cellular pathways in these hiPSC-derived renal cells, and the results were in agreement with human and animal data. Our methods will enable the development of personalized or disease-specific hiPSC-based renal in vitro models for compound screening and nephrotoxicity prediction.

Synthesis of pyrazolylbisindoles over mesoporous Lewis acidic ZrTUD-1: Potential application in selective Cu(2+) colorimetric detection.

A series of pyrazolylbisindole (PBI) derivatives were prepared by simple condensation of indole and pyrazole aldehyde utilizing amorphous mesoporous ZrTUD-1 having predominant Lewis acid sites. The applicability of pyrazolylbisindolyl derivate as a colorimetric chemosensor with high selectivity toward Cu(2+) over other cations were tested. Among heavy and transition metal (HTM) ions in CH3CN solution, the probe only sensed Cu(2+) detectable by naked eye. The sensor exhibited a new absorption band at 488 nm (a red shift of 206 nm from 282 nm) with a large colorimetric response and affinity to Cu(2+) over other cations tested (Al(3+), Pb(2+), Cd(2+), Mg(2+), Mn(2+), Zn(2+), K(+), Fe(2+), Ca(2+), Cu(2+) and Hg(2+)).

A novel pyrazole biscoumarin based chemosensors for the selective detection of Cu(2+) and Zn(2+) ions.

A novel chemosensor based on pyrazole biscoumarin molecule "4-hydroxy-3-((4-hydroxy-2-oxo-2H-chromen-3-yl)(1,3-diphenyl-1H-pyrazol-4-yl)methyl)-2H-chromen-2-one" (PBC) was synthesized by a simple method. The chemosensing properties of PBC towards transition metal ions like Cu(2+) and Zn(2+) by naked eye, UV-Visible and fluorescence spectroscopic methods were described. The PBC solution with Cu(2+) and Zn(2+) ion showed brown and blue colour respectively. The UV-Visible spectra of PBC with Cu(2+) and Zn(2+) ions exposed their corresponding absorption maxima. Further, the Job's plot method confirmed the 1:1 and 2:1 stoichiometry of the complex formation between the PBC with Cu(2+) and Zn(2+) ions respectively. The fluorescence enhancement of PBC on binding with Cu(2+) and Zn(2+) is due to the inhibition of photo induced electron transfer mechanism.

Polysulfone membranes coated with polymerized 3,4-dihydroxy-l-phenylalanine are a versatile and cost-effective synthetic substrate for defined long-term cultures of human pluripotent stem cells.

Clinical and industrial applications of human pluripotent stem cells (hPSC) require large amounts of cells that have been expanded under defined conditions. Labor-intensive techniques and ill-defined or expensive compounds and substrates are not applicable. Here we describe a chemically defined synthetic substrate consisting of polysulfone (PSF) membranes coated with polymerized 3,4-dihydroxy-l-phenylalanine (DOPA). DOPA/PSF is inexpensive and can be easily produced at various shapes and sizes. DOPA/PSF supports long-term self-renewal of undifferentiated human embryonic (hESC) and human induced pluripotent stem cells (hiPSC) under defined conditions. Pluripotency is maintained for at least 10 passages. Adhesion of hPSC to DOPA/PSF is mainly mediated by a specific integrin heterodimer. Proliferation and gene expression patterns on DOPA/PSF and control substrates are comparable. Labor-intensive cultivation methods and use of serum or coating with proteins are not required. Together, these features make DOPA/PSF attractive for applications where large-scale expansion of human pluripotent stem cells under defined conditions is essential.

Identification of nephrotoxic compounds with embryonic stem-cell-derived human renal proximal tubular-like cells.

The kidney is a major target for drug-induced toxicity, and the renal proximal tubule is frequently affected. Nephrotoxicity is typically detected only late during drug development, and the nephrotoxic potential of newly approved drugs is often underestimated. A central problem is the lack of preclinical models with high predictivity. Validated in vitro models for the prediction of nephrotoxicity are not available. Major problems are related to the identification of appropriate cell models and end points. As drug-induced kidney injury is associated with inflammatory reactions, we explored the expression of inflammatory markers as end point for renal in vitro models. In parallel, we developed a new cell model. Here, we combined these approaches and developed an in vitro model with embryonic stem-cell-derived human renal proximal tubular-like cells that uses the expression of interleukin (IL)-6 and IL-8 as end points. The predictivity of the model was evaluated with 41 well-characterized compounds. The results revealed that the model predicts proximal tubular toxicity in humans with high accuracy. In contrast, the predictivity was low when well-established standard in vitro assays were used. Together, the results show that high predictivity can be obtained with in vitro models employing pluripotent stem cell-derived human renal proximal tubular-like cells.

A novel design of bioartificial kidneys with improved cell performance and haemocompatibility.

Treatment with bioartificial kidneys had beneficial effects in animal experiments and improved survival of critically ill patients with acute kidney injury in a Phase II clinical trial. However, a Phase II b clinical trial failed. This and other results suggested various problems with the current design of bioartificial kidneys. We propose a novel design to improve various properties of device, including haemocompatibility and cell performance. An important feature of the novel design is confinement of the blood to the lumina of the hollow fibre membranes. This avoids exposure of the blood to the non-haemocompatible outer surfaces of hollow fibre membranes, which usually occurs in bioartificial kidneys. We use these outer surfaces as substrate for cell growth. Our results show that commercial hollow fibre membranes can be directly applied in the bioreactor when human primary renal proximal tubular cells are grown in this configuration, and no coatings are required for the formation of robust and functional renal epithelia. Furthermore, we demonstrate that the bioreactor unit produces significant amounts of interleukins. This result helps to understand the immunomodulatory effects of bioartificial kidneys, which have been observed previously. The novel bioartificial kidney design outlined here and the results obtained would be expected to improve the safety and performance of bioartificial kidneys and to contribute to a better understanding of their effects.

Human embryonic stem cells differentiate into functional renal proximal tubular-like cells.

Renal cells are used in basic research, disease models, tissue engineering, drug screening, and in vitro toxicology. In order to provide a reliable source of human renal cells, we developed a protocol for the differentiation of human embryonic stem cells into renal epithelial cells. The differentiated stem cells expressed markers characteristic of renal proximal tubular cells and their precursors, whereas markers of other renal cell types were not expressed or expressed at low levels. Marker expression patterns of these differentiated stem cells and in vitro cultivated primary human renal proximal tubular cells were comparable. The differentiated stem cells showed morphological and functional characteristics of renal proximal tubular cells, and generated tubular structures in vitro and in vivo. In addition, the differentiated stem cells contributed in organ cultures for the formation of simple epithelia in the kidney cortex. Bioreactor experiments showed that these cells retained their functional characteristics under conditions as applied in bioartificial kidneys. Thus, our results show that human embryonic stem cells can differentiate into renal proximal tubular-like cells. Our approach would provide a source for human renal proximal tubular cells that are not affected by problems associated with immortalized cell lines or primary cells.

Perinuclear positioning of the inactive human cystic fibrosis gene depends on CTCF, A-type lamins and an active histone deacetylase.

The nuclear positioning of mammalian genes often correlates with their functional state. For instance, the human cystic fibrosis transmembrane conductance regulator (CFTR) gene associates with the nuclear periphery in its inactive state, but occupies interior positions when active. It is not understood how nuclear gene positioning is determined. Here, we investigated trichostatin A (TSA)-induced repositioning of CFTR in order to address molecular mechanisms controlling gene positioning. Treatment with the histone deacetylase (HDAC) inhibitor TSA induced increased histone acetylation and CFTR repositioning towards the interior within 20 min. When CFTR localized in the nuclear interior (either after TSA treatment or when the gene was active) consistent histone H3 hyperacetylation was observed at a CTCF site close to the CFTR promoter. Knockdown experiments revealed that CTCF was essential for perinuclear CFTR positioning and both, CTCF knockdown as well as TSA treatment had similar and CFTR-specific effects on radial positioning. Furthermore, knockdown experiments revealed that also A-type lamins were required for the perinuclear positioning of CFTR. Together, the results showed that CTCF, A-type lamins and an active HDAC were essential for perinuclear positioning of CFTR and these components acted on a CTCF site adjacent to the CFTR promoter. The results are consistent with the idea that CTCF bound close to the CFTR promoter, A-type lamins and an active HDAC form a complex at the nuclear periphery, which becomes disrupted upon inhibition of the HDAC, leading to the observed release of CFTR.

Effects of bone morphogenetic proteins on primary human renal cells and the generation of bone morphogenetic protein-7-expressing cells for application in bioartificial kidneys.

Bioartificial kidneys (BAKs) contain renal cells, and primary human renal proximal tubule cells (HPTCs) have been applied in clinical trials with BAKs. Cell performance within the device is critical. HPTC performance is often compromised under in vitro conditions because of dedifferentiation, transdifferentiation, and tubule formation on substrate surfaces. Herein we tested whether treatments with human recombinant bone morphogenetic protein (BMP)-2 or BMP-7 would improve HPTC performance. We found that both growth factors improved HPTC performance, but more consistent results were obtained with BMP-7. The effects were strongly concentration dependent, and for BMP-7, 25 ng/mL was the optimal concentration, which improved HPTC performance under static and under bioreactor conditions. As an alternative to supplementation with the purified growth factor, we generated HPTCs secreting human recombinant BMP-7. BMP-7 secreted by the cells was bioactive and improved the functional performance of HPTCs, in agreement with our other findings. Together, the results suggested that either supplementation with purified BMP-7 or BMP-7-producing cells could be used to improve cell performance in BAKs. BAKs with BMP-7-producing cells could also be used to deliver the growth factor to kidney patients. Our results suggested that the amount of BMP-7 produced by HPTCs would be sufficient for therapeutic applications.

The use of a library of industrial materials to determine the nature of substrate-dependent performance of primary adherent human cells.

We developed a library of industrial materials, which can be applied to any adherent cell type for determining cell-material interactions. Bulk and surface chemistry as well as other material properties were characterized. The library covered broad ranges of various material properties. We applied the library to primary human endothelial and epithelial cells, which play important roles in tissue engineering and biomedical applications. The results revealed that substrate stiffness was the major determinant of cell performance. The ability to grow and differentiate on stiff or more compliant materials was cell type-dependent, but cell performance was consistently best on stiff and smooth materials. These results give new insights into the nature of substrate-dependent performance of primary human cells and are potentially useful for the development of improved biomaterials. The materials of the library can be easily accessed by the scientific community to determine cell-material interactions of any adherent cell type of interest.

The performance of primary human renal cells in hollow fiber bioreactors for bioartificial kidneys.

Bioartificial kidneys (BAKs) containing human primary renal proximal tubule cells (HPTCs) have been applied in clinical trials. The results were encouraging, but also showed that more research is required. Animal cells or cell lines are not suitable for clinical applications, but have been mainly used in studies on BAK development as large numbers of such cells could be easily obtained. It is difficult to predict HPTC performance based on data obtained with other cell types. To enable more extensive studies on HPTCs, we have developed a bioreactor containing single hollow fiber membranes that requires relatively small amounts of cells. Special hollow fiber membranes with the skin layer on the outer surface and consisting of polyethersulfone/polyvinylpyrrolidone were developed. The results suggested that such hollow fiber membranes were more suitable for the bioreactor unit of BAKs than membranes with an inner skin layer. An HPTC-compatible double coating was applied to the insides of the hollow fiber membranes, which sustained the formation of functional epithelia under bioreactor conditions. Nevertheless, the state of differentiation of the primary human cells remained a critical issue and should be further addressed. The bioreactor system described here will facilitate further studies on the relevant human cell type.

CMOS image sensor for detection of interferon gamma protein interaction as a point-of-care approach.

Complementary metal oxide semiconductor (CMOS)-based image sensors have received increased attention owing to the possibility of incorporating them into portable diagnostic devices. The present research examined the efficiency and sensitivity of a CMOS image sensor for the detection of antigen-antibody interactions involving interferon gamma protein without the aid of expensive instruments. The highest detection sensitivity of about 1 fg/ml primary antibody was achieved simply by a transmission mechanism. When photons are prevented from hitting the sensor surface, a reduction in digital output occurs in which the number of photons hitting the sensor surface is approximately proportional to the digital number. Nanoscale variation in substrate thickness after protein binding can be detected with high sensitivity by the CMOS image sensor. Therefore, this technique can be easily applied to smartphones or any clinical diagnostic devices for the detection of several biological entities, with high impact on the development of point-of-care applications.