Developmental relationship between junctional epithelium and epithelial rests of Malassez.

Keratin 17 (K17) is thought to be a candidate target gene for regulation by Lymphoid Enhancer Factor-1 (Lef-1). K17 is a marker that distinguishes junctional epithelium (JE) from epithelial rests of Malassez (ERM). However, the relationship of Lef-1 to K17 is not clear in this context. Moreover, the expression of other keratins such as K5, K6, K7 and K16 is not reported. Therefore, the aim of our study was to assay the expression of K5, K6, K7, K14, K16, K17 and Lef-1 in postnatal developing teeth, and clarify the corresponding immunophenotypes of the JE and ERM. Upper jaws of Wistar rats aged from postnatal (PN) day 3.5 to PN21 were used and processed for immunohistochemistry. K5 and K14 were intensely expressed in inner enamel epithelium (IEE), reduced enamel epithelium (REE), ERM and JE. There was no staining for K16 in the tissue, except for strong staining in the oral epithelium. Specifically, at PN3.5 and PN7, K17 was initially strongly expressed and then negative in the IEE. At PN16 and PN21, both REE and ERM were strongly stained for K17, whereas K17 was negative in the JE. In addition, K6, K7 and Lef-1 were not detected in any tissue investigated. REE and ERM have an identical keratin expression pattern before eruption, while JE differs from ERM in the expression of K17 after eruption. The expression of K17 does not coincide with that of Lef-1. These data indicate that JE has a unique phenotype different from ERM, which is of odontogenic origin.

Bioenergy production from chicken feather waste by anaerobic digestion and bioelectrochemical systems.

BACKGROUND: Poultry feather waste has a potential for bioenergy production because of its high protein content. This research explored the use of chicken feather hydrolysate for methane and hydrogen production via anaerobic digestion and bioelectrochemical systems, respectively. Solid state fermentation of chicken waste was conducted using a recombinant strain of Bacillus subtilis DB100 (p5.2). RESULTS: In the anaerobic digestion, feather hydrolysate produced maximally 0.67 Nm3 CH4/kg feathers and 0.85 mmol H2/day.L concomitant to COD removal of 86% and 93%, respectively. The bioelectrochemical systems used were microbial fuel and electrolysis cells. In the first using a microbial fuel cell, feather hydrolysate produced electricity with a maximum cell potential of 375 mV and a current of 0.52 mA. In the microbial electrolysis cell, the hydrolysate enhanced the hydrogen production rate to 7.5 mmol/day.L, with a current density of 11.5 A/m2 and a power density of 9.26 W/m2. CONCLUSIONS: The data indicated that the sustainable utilization of keratin hydrolysate to produce electricity and biohydrogen via bioelectrical chemical systems is feasible. Keratin hydrolysate can produce electricity and biofuels through an integrated aerobic-anaerobic fermentation system.

Unraveling UVB effects: Catalase activity and molecular alterations in the stratum corneum.

AIM: Ultraviolet B (UVB) radiation can compromise the functionality of the skin barrier through various mechanisms. We hypothesize that UVB induce photochemical alterations in the components of the outermost layer of the skin, known as the stratum corneum (SC), and modulate its antioxidative defense mechanisms. Catalase is a well-known antioxidative enzyme found in the SC where it acts to scavenge reactive oxygen species. However, a detailed characterization of acute UVB exposure on the activity of native catalase in the SC is lacking. Moreover, the effects of UVB irradiation on the molecular dynamics and organization of the SC keratin and lipid components remain unclear. Thus, the aim of this work is to characterize consequences of UVB exposure on the structural and antioxidative properties of catalase, as well as on the molecular and global properties of the SC matrix surrounding the enzyme. EXPERIMENTS: The effect of UVB irradiation on the catalase function is investigated by chronoamperometry with a skin covered oxygen electrode, which probes the activity of native catalase in the SC matrix. Circular dichroism is used to explore changes of the catalase secondary structure, and gel electrophoresis is used to detect fragmentation of the enzyme following the UVB exposure. UVB induced alterations of the SC molecular dynamics and structural features of the SC barrier, as well as its water sorption behavior, are investigated by a complementary set of techniques, including natural abundance 13C polarization transfer solid-state NMR, wide-angle X-ray diffraction, Fourier transform infrared (FTIR) spectroscopy, and dynamic vapor sorption microbalance. FINDINGS: The findings show that UVB exposure impairs the antioxidative function of catalase by deactivating both native catalase in the SC matrix and lyophilized catalase. However, UVB radiation does not alter the secondary structure of the catalase nor induce any observable enzyme fragmentation, which otherwise could explain deactivation of its function. NMR measurements on SC samples show a subtle increase in the molecular mobility of the terminal segments of the SC lipids, accompanied by a decrease in the mobility of lipid chain trans-gauche conformers after high doses of UVB exposure. At the same time, the NMR data suggest increased rigidity of the polypeptide backbone of the keratin filaments, while the molecular mobility of amino acid residues in random coil domains of keratin remain unaffected by UVB irradiation. The FTIR data show a consistent decrease in absorbance associated with lipid bond vibrations, relative to the main protein bands. Collectively, the NMR and FTIR data suggest a small modification in the composition of fluid and solid phases of the SC lipid and protein components after UVB exposure, unrelated to the hydration capacity of the SC tissue. To conclude, UVB deactivation of catalase is anticipated to elevate oxidative stress of the SC, which, when coupled with subtle changes in the molecular characteristics of the SC, may compromise the overall skin health and elevate the likelihood of developing skin disorders.

Keratin Formed Bioadhesive Ophthalmic Gel for the Bacterial Conjunctivitis Treatment.

Keratin has the potential to function as the gel matrix in an ophthalmic formulation for the encapsulation of the macrolide antibiotic azithromycin. The quality of this formulation was thoroughly evaluated through various analyses, such as in vitro release assessment, rheological examination, intraocular retention studies in rabbits, assessment of bacteriostatic efficacy, and safety evaluations. It is worth mentioning that the gel demonstrated shear thinning properties and exhibited characteristics of an elastic solid, thereby confirming its structural stability. The gel demonstrated a notable affinity for mucosal surfaces in comparison to traditional azithromycin aqueous solutions. In vitro release testing revealed that drug release transpired via diffusion mechanisms, following a first-order kinetic release pattern. Additionally, the formulated gel exhibited remarkable antibacterial efficacy against Staphylococcus aureus and Pseudomonas aeruginosa in bacteriostatic evaluations. Lastly, safety assessments confirmed that the gel eye drops induced minimal irritation and displayed no apparent cytotoxicity, indicating their good safety and biocompatibility for ocular application. Thus, these findings indicated that the prepared azithromycin gel eye drops complied with the requisite standards for ophthalmic preparations.

Wound Dressing Based on Silver Nanoparticle Embedded Wool Keratin Electrospun Nanofibers Deposited on Cotton Fabric: Preparation, Characterization, Antimicrobial Activity, and Cytocompatibility.

Wool keratin (WK) protein is attractive for wound dressing and biomedical applications due to its excellent biodegradability, cytocompatibility, and wound-healing properties. In this work, WK-based wound dressings were prepared by depositing WK/poly(vinyl alcohol) (PVA) and silver nanoparticle (Ag NP)-embedded WK/PVA composite nanofibrous membranes on cotton fabrics by electrospinning. Ag NPs were biosynthesized by reduction and stabilization with sodium alginate. The formed Ag NPs were characterized by ultraviolet-visible and Fourier transform infrared (FTIR) spectroscopy, and their size was determined by transmission electron microscopy and image analysis. The formed Ag NPs were spherical and had an average diameter of 9.95 nm. The produced Ag NP-embedded WK/PVA composite nanofiber-deposited cotton fabric surface was characterized by FTIR and dynamic contact angle measurements, and the nanofiber morphologies were characterized by scanning electron microscopy. The average diameter of the nanofibers formed by 0.1% Ag NP-embedded WK/PVA solution was 146.7 nm. The antibacterial activity of the surface of cotton fabrics coated with electrospun composite nanofibers was evaluated against the two most common wound-causing pathogens, Staphylococcus aureus and Pseudomonas aeruginosa. The cotton fabric coated with 0.1% Ag NP-embedded WK/PVA nanofibers showed very good antibacterial activity against both pathogens, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay results showed good cytocompatibility against L-929 mouse fibroblast cells. However, the increase in Ag NP content in the nanofibers to 0.2% negatively affected the cell viability due to the high release rate of Ag ions. The results achieved show that the developed wound dressing has good potential for wound healing applications.

In vitro generation of epidermal keratinocytes from human CD34-positive hematopoietic stem cells.

The epidermis is largely composed of keratinocytes (KCs), and the proliferation and differentiation of KCs from the stratum basale to the stratum corneum is the cellular hierarchy present in the epidermis. In this study, we explore the differentiation abilities of human hematopoietic stem cells (HSCs) into KCs. Cultured HSCs positive for CD34, CD45, and CD133 with prominent telomerase activity were induced with keratinocyte differentiation medium (KDM), which is composed of bovine pituitary extract (BPE), epidermal growth factor (EGF), insulin, hydrocortisone, epinephrine, transferrin, calcium chloride (CaCl2), bone morphogenetic protein 4 (BMP4), and retinoic acid (RA). Differentiation was monitored through the expression of cytokeratin markers K5 (keratin 5), K14 (keratin 14), K10 (keratin 10), K1 (keratin 1), transglutaminase 1 (TGM1), involucrin (IVL), and filaggrin (FLG) on day 0 (D0), day 6 (D6), day 11 (D11), day 18 (D18), day 24 (D24), and day 30 (D30) using immunocytochemistry, fluorescence microscopy, flow cytometry, qPCR, and Western blotting. The results revealed the expression of K5 and K14 genes in D6 cells (early keratinocytes), K10 and K1 genes in D11-D18 cells (mature keratinocytes) with active telomerase enzyme, and FLG, IVL, and TGM1 in D18-D24 cells (terminal keratinocytes), and by D30, the KCs were completely enucleated similar to cornified matrix. This method of differentiation of HSCs to KCs explains the cellular order exists in the normal epidermis and opens the possibility of exploring the use of human HSCs in the epidermal differentiation.

Peptide recovery from chicken feather keratin and their anti-biofilm properties against methicillin-resistant Staphylococcus aureus (MRSA).

Bacteria have the potential to adhere to abiotic surfaces, which has an undesirable effect in the food industry because they can survive for sustained periods through biofilm formation. In this study, an antibacterial peptide (ABP), with a molecular mass of 3861 Da, was purified from hydrolyzed chicken feathers using a locally isolated keratinolytic bacterium, namely Rhodococcus erythropolis, and its antibacterial and antibiofilm potential were investigated against planktonic and biofilm cells of Methicillin-Resistant Staphylococcus Aureus (MRSA). The results demonstrated that purified ABP showed the growth inhibition of MRSA cells with the minimum inhibitory concentration (MIC) of 45 µg/ml and disrupted MRSA biofilm formation at a concentration of 200 ug/ml, which results were confirmed by scanning electron micrograph (SEM). Moreover, the secondary structures of the peptide were assessed as part of the FTIR analysis to evaluate its mode of action. ExPASy tools were used to predict the ABP sequence, EPCVQUQDSRVVIQPSPVVVVTLPGPILSSFPQNTA, from a chicken feather keratin sequence database following in silico digestion by trypsin. Also, ABP had 54.29% hydrophobic amino acids, potentially contributing to its antimicrobial activity. The findings of toxicity prediction of the peptide by the ToxinPred tool revealed that ABP had non-toxic effects. Thus, these results support the potential of this peptide to be used as an antimicrobial agent for the treatment or prevention of MRSA biofilm formation in feed, food, or pharmaceutical applications.

Throwing off the keratin chains: a potential therapy for hereditary podocytopathy.

In the current issue, Kuzmuk et al. offer a therapeutic option for patients with NPHS2 R138Q-associated nephrotic syndrome. For the first time in hereditary podocytopathies, this is offered by restoring the membrane targeting of a pathogenic protein. The idea that it is enough to liberate podocin from the trap of keratin 8, a key member of endoplasmic-reticulum-associated protein degradation complex, was brilliantly recognized based on former results obtained in cystic fibrosis.

Rhizoctonia solani disease suppression: addition of keratin-rich soil amendment leads to functional shifts in soil microbial communities.

Promoting soil suppressiveness against soil borne pathogens could be a promising strategy to manage crop diseases. One way to increase the suppression potential in agricultural soils is via the addition of organic amendments. This microbe-mediated phenomenon, although not fully understood, prompted our study to explore the microbial taxa and functional properties associated with Rhizoctonia solani disease suppression in sugar beet seedlings after amending soil with a keratin-rich waste stream. Soil samples were analyzed using shotgun metagenomics sequencing. Results showed that both amended soils were enriched in bacterial families found in disease suppressive soils before, indicating that the amendment of keratin-rich material can support the transformation into a suppressive soil. On a functional level, genes encoding keratinolytic enzymes were found to be abundant in the keratin-amended samples. Proteins enriched in amended soils were those potentially involved in the production of secondary metabolites/antibiotics, motility, keratin-degradation, and contractile secretion system proteins. We hypothesize these taxa contribute to the amendment-induced suppression effect due to their genomic potential to produce antibiotics, secrete effectors via the contractile secretion system, and degrade oxalate-a potential virulence factor of R. solani-while simultaneously possessing the ability to metabolize keratin.

Rational engineering S1' substrate binding pocket to enhance substrate specificity and catalytic activity of thermal-stable keratinase for efficient keratin degradation.

This study reports the rational engineering of the S1' substrate-binding pocket of a thermally-stable keratinase from Pseudomonas aeruginosa 4-3 (4-3Ker) to improve substrate specificity to typical keratinase (K/C > 0.5) and catalytic activity without compromising thermal stability for efficient keratin degradation. Of 10 chosen mutation hotspots in the S1' substrate-binding pocket, the top three mutations M128R, A138V, and V142I showing the best catalytic activity and substrate specificity were identified. Their double and triple combinatorial mutants synergistically overcame limitations of single mutants, fabricating an excellent M128R/A138V/V142I triple mutant which displayed a 1.21-fold increase in keratin catalytic activity, 1.10-fold enhancement in keratin/casein activity ratio, and a 3.13 °C increase in half-inactivation temperature compared to 4-3Ker. Molecular dynamics simulations revealed enhanced flexibility of critical amino acid residues at the substrate access tunnel, improved global protein rigidity, and heightened hydrophobicity within the active site likely underpinned the increased catalytic activity and substrate specificity. Additionally, the triple mutant improved the feather degradation rate by 32.86 % over the wild-type, far exceeding commercial keratinase in substrate specificity and thermal stability. This study exemplified engineering a typical keratinase with enhanced substrate specificity, catalytic activity, and thermal stability from thermally-stable 4-3Ker, providing a more robust tool for feather degradation.

Keratin-based bioplastics extracted from chicken feathers: Effect of chitosan concentration on the structural, chemical bonding, and mechanical properties of bioplastics.

Keratin was synthesized by alkaline hydrolysis from chicken feathers and then continue by casting method for producing bioplastics with additional various amounts of chitosan as a filler, polyvinyl alcohol (PVA) and glycerol as a plasticizer. The main purpose is analysis the effect of chitosan on the structural properties using quantitative analysis of X-ray diffraction (XRD) spectra, chemical bonding by Fourier transforms infrared (FTIR) spectra, and mechanical properties by texture analyser to the keratin-based bioplastics. Biodegradation of bioplastics was analysed from the loss of weight by burying in the soil. It's found that, the additional of chitosan (0 %, 2 %, 5 %, and 8 %) increased the crystallinity of bioplastics by 11.83 %, 11.12 %, 18.99 %, and 17.03 %, respectively, but decreasing tensile strength and elasticity of bioplastics. Degradation of bioplastic keratin-based shows that the addition of chitosan can reduce the degradation time which is directly proportional to the loss of CO bonds. The highest degradation rate is 89.29 % in 49 days for keratin-based bioplastics with 8 % chitosan, indicated that high potential for future production.

EBS in Children with De Novo Pathogenic Variants Disturbing Krt14.

Epidermolysis bullosa simplex (EBS) is a dermatological condition marked by skin fragility and blister formation resulting from separation within the basal layer of the epidermis, which can be attributed to various genetic etiologies. This study presents three pathogenic de novo variants in young children, with clinical manifestations appearing as early as the neonatal period. The variants contribute to the EBS phenotype through two distinct mechanisms: direct keratin abnormalities due to pathogenic variants in the Krt14 gene, and indirect effects via pathogenic mutation in the KLHL24 gene, which interfere with the natural proteasome-mediated degradation pathway of KRT14. We report one severe case of EBS with mottled pigmentation arising from the Met119Thr pathogenic variant in KRT14, another case involving a pathogenic KLHL24 Met1Val variant, and a third case featuring the hot spot mutation Arg125His in KRT14, all manifesting within the first few weeks of life. This research underscores the complexity of genetic influences in EBS and highlights the importance of early genetic screening for accurate diagnosis and management.

Evolutionary origin of Hoxc13-dependent skin appendages in amphibians.

Cornified skin appendages, such as hair and nails, are major evolutionary innovations of terrestrial vertebrates. Human hair and nails consist largely of special intermediate filament proteins, known as hair keratins, which are expressed under the control of the transcription factor Hoxc13. Here, we show that the cornified claws of Xenopus frogs contain homologs of hair keratins and the genes encoding these keratins are flanked by promoters in which binding sites of Hoxc13 are conserved. Furthermore, these keratins and Hoxc13 are co-expressed in the claw-forming epithelium of frog toe tips. Upon deletion of hoxc13, the expression of hair keratin homologs is abolished and the development of cornified claws is abrogated in X. tropicalis. These results indicate that Hoxc13-dependent expression of hair keratin homologs evolved already in stem tetrapods, presumably as a mechanism for protecting toe tips, and that this ancestral genetic program was coopted to the growth of hair in mammals.

Thermoresponsive keratin-methylcellulose self-healing injectable hydrogel accelerating full-thickness wound healing by promoting rapid epithelialization.

Chronic wounds suffer from impaired healing due to microbial attack and poor vascular growth. Thermoresponsive hydrogels gained attention in wound dressing owing to their gelation at physiological temperature enabling them to take the shape of asymmetric wounds. The present study delineates the development of thermoresponsive hydrogel (MCK), from hair-derived keratin (K) and methylcellulose (MC) in the presence of sodium sulfate. The gelation temperature (Tg) of this hydrogel is in the range of 30 °C to 33 °C. Protein-polymer interaction leading to thermoreversible sol-gel transition involved in MCK blends has been analyzed and confirmed by FTIR, XRD, and thermal studies. Keratin, has introduced antioxidant properties to the hydrogel imparted cytocompatibility towards human dermal fibroblasts (HDFs) as evidenced by both MTT and live dead assays. In vitro wound healing assessment has been shown by enhanced migration of HDFs in the presence of MCK hydrogel compared to the control. Also, CAM assay and CD31 expression by the Wistar rat model has shown increased blood vessel branching after the implantation of MCK hydrogel. Further, in vivo study, demonstrated MCK efficacy of hydrogel in accelerating full-thickness wounds with minimal scarring in Wistar rats, re-epithelialization, and reinstatement of the epidermal-dermal junction thereby exhibiting clinical relevance for chronic wounds.

A facile method to fabricate versatile keratin cryogels for tissue engineering applications.

Human hair keratin (HHK) has been extensively explored as a biomaterial for soft tissue regeneration due to their excellent bioactivity and biocompatibility. The possibility to fabricate HHK into three-dimensional (3D) hydrogels with physical properties resembling soft tissues has been well demonstrated. However, conventional keratin hydrogels often exhibit a dense architecture that could hinder cell filtration. In the present study, HHK-based cryogels were fabricated using a freeze-thaw (FT) method, where oxidized dopamine (ODA) was employed to covalently crosslink thiol/amine rich-keratin molecules at sub-zero temperatures. The obtained HHK-ODA cryogels have micron-sized pores ranging between 100 and 200 µm and mechanical properties that can be tuned by varying the crosslinking density between ODA and HHK. Through optimization of the weight content of ODA and the number of FT cycles, the compressive strengths and stiffnesses of these cryogels achieved 15-fold increments from ∼1.5 kPa to ∼22 kPa and ∼300 Pa to ∼5000 Pa, respectively. The HHK-ODA cryogels competently supported human dermal fibroblast spreading and proliferation. Overall, this study exhibited a facile method to fabricate mechanically superior keratin-based cryogels with cell compatible microarchitecture, circumventing the need for complicated chemical modifications and the use of cytotoxic crosslinkers.

Keratin-containing scaffolds for tissue engineering applications: a review.

In tissue engineering and regenerative medicine applications, the utilization of bioactive materials has become a routine tool. The goal of tissue engineering is to create new organs and tissues by combining cell biology, materials science, reactor engineering, and clinical research. As part of the growth pattern for primary cells in an organ, backing material is frequently used as a supporting material. A porous three-dimensional (3D) scaffold can provide cells with optimal conditions for proliferating, migrating, differentiating, and functioning as a framework. Optimizing the scaffolds' structure and altering their surface may improve cell adhesion and proliferation. A keratin-based biomaterials platform has been developed as a result of discoveries made over the past century in the extraction, purification, and characterization of keratin proteins from hair and wool fibers. Biocompatibility, biodegradability, intrinsic biological activity, and cellular binding motifs make keratin an attractive biomaterial for tissue engineering scaffolds. Scaffolds for tissue engineering have been developed from extracted keratin proteins because of their capacity to self-assemble and polymerize into intricate 3D structures. In this review article, applications of keratin-based scaffolds in different tissues including bone, skin, nerve, and vascular are explained based on common methods of fabrication such as electrospinning, freeze-drying process, and sponge replication method.

Interaction of human dendritic cell receptor DEC205/CD205 with keratins.

DEC205 (CD205) is one of the major endocytic receptors on dendritic cells and has been widely used as a receptor target in immune therapies. It has been shown that DEC205 can recognize dead cells through keratins in a pH-dependent manner. However, the mechanism underlying the interaction between DEC205 and keratins remains unclear. Here we determine the crystal structures of an N-terminal fragment of human DEC205 (CysR∼CTLD3). The structural data show that DEC205 shares similar overall features with the other mannose receptor family members such as the mannose receptor and Endo180, but the individual domains of DEC205 in the crystal structure exhibit distinct structural features that may lead to specific ligand binding properties of the molecule. Among them, CTLD3 of DEC205 adopts a unique fold of CTLD, which may correlate with the binding of keratins. Furthermore, we examine the interaction of DEC205 with keratins by mutagenesis and biochemical assays based on the structural information and identify an XGGGX motif on keratins that can be recognized by DEC205, thereby providing insights into the interaction between DEC205 and keratins. Overall, these findings not only improve the understanding of the diverse ligand specificities of the mannose receptor family members at the molecular level but may also give clues for the interactions of keratins with their binding partners in the corresponding pathways.

Core-shell nanofibers containing L-arginine stimulates angiogenesis and full thickness dermal wound repair.

Despite the advances in medicine, wound healing is still challenging and piques the interest of biomedical engineers to design effective wound dressings using natural and artificial polymers. In present study, coaxial electrospinning was employed to fabricate core-shell nanofiber-based wound dressing, with core composed of polyacrylamide (PAAm) and shell comprising 0.5 % solution of L-Arginine (L-Arg) in aloe vera and keratin (AloKr). Aloe vera and keratin were added as natural polymers to promote angiogenesis, reduce inflammation, and provide antibacterial activity, whereas PAAm in core was used to improve the tensile properties of the wound dressing. Moreover, L-Arg was incorporated in shell to promote angiogenesis and collagen synthesis. The fiber diameter of PAAm/(AloKr/L-Arg) core-shell fibers was (93.33 ± 35.11 nm) with finer and straighter fibers and higher water holding capacity due to increased surface area to volume ratio. In terms of tensile properties, the PAAm/(AloKr/L-Arg) core-shell nanofibers with tensile strength and elastic modulus of 2.84 ± 0.27 MPa and 62.15 ± 5.32 MPa, respectively, showed the best mechanical performance compared to other nanofibers tested. Furthermore, PAAm/(AloKr/L-Arg) exhibited the highest L-Arg release (87.62 ± 3.02 %) and viability of L929 cells in vitro compared to other groups. In addition, the highest rate of in vivo full thickness wound healing was observed in PAAm/(AloKr/L-Arg) group compared to other groups. It significantly enhanced the angiogenesis, neovascularization, and cell proliferation. The prepared PAAm/(AloKr/L-Arg) core-shell nanofibrous dressing could be promising for full-thickness wound healing.

Nanotoxicity of two-dimensional nanomaterials on human skin and the structural evolution of keratin protein.

Two-dimensional (2D) materials have been increasingly widely used in biomedical and cosmetical products nowadays, yet their safe usage in human body and environment necessitates a comprehensive understanding of their nanotoxicity. In this work, the effect of pristine graphene and graphene oxide (GO) on the adsorption and conformational changes of skin keratin using molecular dynamics simulations. It is found that skin keratin can be absorbed through various noncovalent driving forces, such as van der Waals (vdW) and electrostatics. In the case of GO, the oxygen-containing groups prevent tighter contact between skin keratin and the graphene basal plane through steric effects and electrostatic repulsion. On the other hand, electrostatic attraction and hydrogen bonding enhance their binding affinity to positively charged residues such as lysine and arginine. The secondary structure of skin keratin is better preserved in GO system, suggesting that GO has good biocompatibility. The charged groups on GO surface perform as the hydrogen bond acceptors, which is like to the natural receptors of keratin in this physiological environment. This work contributes to a better knowledge of the nanotoxicity of cutting-edge 2D materials on human health, thereby advancing their potential biological applications.

JAK/STAT3 represents a therapeutic target for colorectal cancer patients with stromal-rich tumors.

Colorectal cancer (CRC) is a heterogenous malignancy underpinned by dysregulation of cellular signaling pathways. Previous literature has implicated aberrant JAK/STAT3 signal transduction in the development and progression of solid tumors. In this study we investigate the effectiveness of inhibiting JAK/STAT3 in diverse CRC models, establish in which contexts high pathway expression is prognostic and perform in depth analysis underlying phenotypes. In this study we investigated the use of JAK inhibitors for anti-cancer activity in CRC cell lines, mouse model organoids and patient-derived organoids. Immunohistochemical staining of the TransSCOT clinical trial cohort, and 2 independent large retrospective CRC patient cohorts was performed to assess the prognostic value of JAK/STAT3 expression. We performed mutational profiling, bulk RNASeq and NanoString GeoMx® spatial transcriptomics to unravel the underlying biology of aberrant signaling. Inhibition of signal transduction with JAK1/2 but not JAK2/3 inhibitors reduced cell viability in CRC cell lines, mouse, and patient derived organoids (PDOs). In PDOs, reduced Ki67 expression was observed post-treatment. A highly significant association between high JAK/STAT3 expression within tumor cells and reduced cancer-specific survival in patients with high stromal invasion (TSPhigh) was identified across 3 independent CRC patient cohorts, including the TrasnSCOT clinical trial cohort. Patients with high phosphorylated STAT3 (pSTAT3) within the TSPhigh group had higher influx of CD66b + cells and higher tumoral expression of PDL1. Bulk RNAseq of full section tumors showed enrichment of NFκB signaling and hypoxia in these cases. Spatial deconvolution through GeoMx® demonstrated higher expression of checkpoint and hypoxia-associated genes in the tumor (pan-cytokeratin positive) regions, and reduced lymphocyte receptor signaling in the TME (pan-cytokeratin- and αSMA-) and αSMA (pan-cytokeratin- and αSMA +) areas. Non-classical fibroblast signatures were detected across αSMA + regions in cases with high pSTAT3. Therefore, in this study we have shown that inhibition of JAK/STAT3 represents a promising therapeutic strategy for patients with stromal-rich CRC tumors. High expression of JAK/STAT3 proteins within both tumor and stromal cells predicts poor outcomes in CRC, and aberrant signaling is associated with distinct spatially-dependant differential gene expression.