Artificial nerve graft constructed by coculture of activated Schwann cells and human hair keratin for repair of peripheral nerve defects

Studies have shown that human hair keratin(HHK) has no antigenicity and excellent mechanical properties. Schwann cells, as unique glial cells in the peripheral nervous system, can be induced by interleukin-1β to secrete nerve growth factor, which promotes neural regeneration. Therefore, HHK with Schwann cells may be a more effective approach to repair nerve defects than HHK without Schwann cells. In this study, we established an artificial nerve graft by loading an HHK skeleton with activated Schwann cells. We found that the longitudinal HHK microfilament structure provided adhesion medium, space and direction for Schwann cells, and promoted Schwann cell growth and nerve fiber regeneration. In addition, interleukin-1β not only activates Schwann cells, but also strengthens their activity and increases the expression of nerve growth factors. Activated Schwann cells activate macrophages, and activated macrophages secrete interleukin-1β, which maintains the activity of Schwann cells. Thus, a beneficial cycle forms and promotes nerve repair. Furthermore, our studies have found that the newly constructed artificial nerve graft promotes the improvements in nerve conduction function and motor function in rats with sciatic nerve injury, and increases the expression of nerve injury repair factors fibroblast growth factor 2 and human transforming growth factor B receptor 2. These findings suggest that this artificial nerve graft effectively repairs peripheral nerve injury.

Close-loop recyclable and flexible halide perovskite@wool keratin sensor with piezoelectric property

Halide perovskites with excellent piezoelectric properties,but their poor stability hinders their largescale application.Herein,a sandwich-structured halide perovskite flexible sensor with good stability was developed according to a three-step procedure as follows:(ⅰ) in-situ growth of wool keratinCsPbBr_(3)(WK-CsPbBr_(3)) using wool keratin in interfacial passivation and coating,(ⅱ) electrospinning of a wool keratin-CsPbBr_(3)/polyacrylonitrile(WCP) nanofiber film,and(ⅲ) coating of the WCP nanofiber with polydimethylsiloxane(PDMS) to obtain a sensor(WCPP).The sensor could generate a piezoelectric voltage of 7.8 V at a pressure of 6 kPa in the stages of pressing and releasing,and the output characteristics did not decline even after 10,000 cycles.Compared to the 4-month stability of the perovskite sensor,WCPP sensor exhibited the output performance even after 16 months,which indicated that wool keratin as a multidentate improved the stability of the halide perovskite.Additionally,the sensor displayed a self-cleaning property and could also light up 14 commercial LEDs.The close-loop recycling of the lead halide perovskite was achieved by dissolving the WCP nanofiber film in DMF and then reelectrospinning.Therefore,the method proposed is a step forward for achieving the commercialization of WK-CsPbBr_(3) and providing new avenues for further utilization of wool waste.

Isolation,Screening and Primary Identification of a Keratin-degrading Fungus

[Objective] The paper was to provide new germplasm sources for efficient and economical degradation and utilization of animal keratin.[Method] The keratin-degrading fungus was isolated,screened and primarily identified by using the combination method of traditional isolation and screening,solid culture-medium degradation and animal test.[Result] A strain of non-pathogenic filamentous fungi with high degradation efficiency was obtained,which was preliminarily identified to be a species in Mucoraceae.[Conclusion] The discovery of the strain enriched the family members of keratin-degrading fungus,and provided new germplasm resources for degradation and utilization of animal keratin.

Tailoring the Meso-Structure of Gold Nanoparticles in Keratin-Based Activated Carbon Toward High-Performance Flexible Sensor

Flexible biosensors with high accuracy and reliable operation in detecting pH and uric acid levels in body fluids are fabricated using well-engineered metaldoped porous carbon as electrode material.The gold nanoparticles@N-doped carbon in situ are prepared using wool keratin as both a novel carbon precursor and a stabilizer.The conducting electrode material is fabricated at 500℃ under customized parameters,which mimics A-B type(two different repeating units) polymeric material and displays excellent deprotonation performance(pH sensitivity).The obtained pH sensor exhibits high pH sensitivity of 57 mV/pH unit and insignificant relative standard deviation of 0.088%.Conversely,the composite carbon material with sp^2 structure prepared at 700℃ is doped with nitrogen and gold nanoparticles,which exhibits good conductivity and electrocatalytic activity for uric acid oxidation.The uric acid sensor has linear response over a range of 1-150 μM and a limit of detection 0.1 μM.These results will provide new avenues where biological material will be the best start,which can be useful to target contradictory applications through molecular engineering at mesoscale.

The structure,tensile properties and water resistance of hydrolyzed feather keratin-based bioplastics

Feather, as a by-product of the poultry industry, has long been treated as a solid waste, which causes environ- mental and economic problems. In this work, the hydrolyzed feather keratin （HFK） was extracted from the chicken feather using a cost-effective method of alkali-extraction and acid-precipitation by applying urea and sodium sulfide. The aim was development and characterization of the eco-friendly films based on the HFK with variable glycerol contents by a thermoplastic process. The thermal analysis showed that high temperature and high pres- sure improved the compatibility between the glycerol and the HFI（ molecules. Also it was shown that the addi- tion of water is necessary in the hot-pressing process of films, The FT-IR analysis indicated that the formation of the new hydrogen bonds between HFK and glycerol. By increasing the glycerol content, the film tensile strength （orb ） decreases from 10,5 MPa to 5.7 MPa and the solubility increases from 15.3% to 20.9%, while the elongation at break （εb） achieves the maximum value of 63,8% for the film with 35% glycerol. The swelling was just below 16.9% at 25 ℃ for 24 h, suggesting a good stability of the films in water. The water vapor permeability （WVP） varied between 3.02 x 10 ^10g. m 2. s-1 . pa-1 and 4.11 x 10-10g · m-2 · s-1 · Pa-1 for the films with 20%and 40% glycerol, respectively. The HFK film was uniform, translucent and tough, which could be used in packaging and agricultural field.

Robust Waterborne Polyurethane/Wool Keratin/Silk Sericin Freeze-Drying Composite Membrane for Heavy Metal Ions Adsorption

Wool keratin(WK)and silk sericin(SS)have the ability to interact with metal ions.In order to take advantage of this potential,a novel environmentally friendly waterborne polyurethane(WPU)/WK/SS membrane named as WPU&WK&SS membrane with crosslinked structure was constructed by freeze-drying via self-assembly style.Surface morphology and chemical structure characterization were investigated by scanning electron microscopy(SEM)and Fourier transform infrared spectroscopy(FT-IR).In addition,the adsorption experiments of Cu2+and Cr6+were performed to evaluate the adsorption of WPU&WK&SS membrane,including adsorption kinetics,adsorption isotherm models and various factors affecting adsorption.Further investigation indicates that the maximum adsorption capacity of Cu2+and Cr6+can reach 54.21 mg·g-1and 85.21 mg·g-1,respectively,which are higher than most of the reported adsorbents.Through adsorption kinetics and thermodynamic analysis,it is find that the pseudo-second-order kinetic equation and the Langmuir adsorption isotherm model are more suitable for the static adsorption of Cu2+and Cr6+by WPU&WK&SS membrane.

Degradation of human hair keratin scaffold material used to repair injured skeletal muscles of rabbits

Objective:To explore the mechanism of the degradation of human hair keratin (HHK) scaffold material implanted in damaged skeletal muscle tissues. Methods: Six New Zealand rabbits with HHK scaffold material implants (composed of 3 different types of HHK material with varied degradation speed) after musclectomy were divided into 3 groups (2 in each group) to observe the degradation of the material at 1, 3, 6weeks after operation. Another rabbit without operation was used as the control group. The degradation of HHK was observed with light microscopy, histochemistry of ubiquitin and electron microscopy. Results:Light microscopy showed that human hair cuticles fell off from the HHK material and emerged, and the macrophagocytes and multinucleate giant cells were attached onto the surface of the material, which became homogeneous at the first postoperative week. The HHK scaffold material was degraded into particles that was phagocytosed by macrophagocytes and multinucleate giant cells at the third week. Ubiquitin enzymatic histochemistry showed that the macrophagocytes and the multinucleate giant cells were positive at the first week. Under electron microscope, HHK scaffold material was degraded into particles, and at the sixth week,part of HHK scaffold material was further degraded. Conclusion: Large mass of the HHK scaffold material is degraded via ubiquitin system, and the resultant particles are phagocytosed and degraded with the cooperation of lysosome and ubiquitin.

Peripheral nerve repair by conduits made of human hair keratin: An experimental study

Objective: To explore the method to repair injured peripheral nerve using conduits made of human hair keratin (HHK). Methods: The tibial nerves of rabbits were transected leaving a gap 10 mm in length between the 2 severed ends, which were either routinely sutured or bridged using HHK nerve conduits. Electro-physiological , anatomical and histological examinations were performed at different time postoperatively. Results: Electrophysiological study showed more obvious improvement in the neural function recovery in rabbits with HHK conduits bridging as compared with that in rabbits with routine suture. In the former group, HHK conduits were gradually degraded and absorbed with large amount of myelinated nerve fibers and Schwann cells regenerated around HHK conduits. In the latter group, however, the nerve tissues around the suture were degenerated and replaced by connective tissues. Conclusion: HHK may induce the regeneration of the nerve fibers and provides an ideal approach to repair nerve damages.

A mutation in the type Ⅱ hair keratin KRT86 gene in a Han family with monilethrix

Monilethrix, a congenital disease of hair, is usually associated with mutations in keratin genes, like KRT81, KRT83 and KRT86. We conducted this study to investigate the mutation of type Ⅱ human basic hair keratin hHb/ KRT gene in a Han family with monilethrix and obtain information for potential pathogenic mechanism study of monilethrix. Peripheral blood samples were drawn for genomic DNA detection. Exon 1 and exon 7 of the KRT81, KRT83 and KRT86 genes were amplified by PCR. All PCR products were sequenced directly using an ABI 310 DNA sequencer. These sequences were aligned with the standard sequences in GenBank using the BLAST software. PCR products were digested with restriction endonuclease and restriction fragment length polymorphism （RFLP） analysis was performed. In this study, we identified one novel mutation, which is a heterozygous transitional mutation of G→A at position 1,289 in exon 7 of the KRT86 gene [R430Q （KRT86）]. RFLP assays for the novel mutation excluded the possibility of polymorphism. The R430Q mutation of the KRT86 gene may be pathogenic for monilethrix. Meanwhile, we did not find any novel mutation or recurrent mutation in exons 1 and 7 of KRT81 and KRT83 and exon 1 of KRT86. There is a potential pathogenic gene in the subjects and our results expand the spectrum of mutations in the hHb6 gene.

Keratinase production and keratin degradation by a mutant strain of Bacillus subtilis

A new feather-degrading bacterium was isolated from a local feather waste site and identified as Bacillus subtilis based on morphological, physiochemical, and phylogenetic characteristics. Screening for mutants with elevated keratinolytic activity using N-methyl-N＇-nitro-N-nitrosoguanidine mutagenesis resulted in a mutant strain KD-N2 producing keratinolytic activity about 2.5 times that of the wild-type strain. The mutant strain produced inducible keratinase in different substrates of feathers, hair, wool and silk under submerged cultivation. Scanning electron microscopy studies showed the degradation of feathers, hair and silk by the keratinase. The optimal conditions for keratinase production include initial pH of 7.5, inoculum size of 2% （v/v）, age of inoculum of 16 h, and cultivation at 23 ℃. The maximum keratinolytic activity of KD-N2 was achieved after 30 h. Essential amino acids like threonine, valine, methionine as well as ammonia were produced when feathers were used as substrates. Strain KD-N2, therefore, shows great promise of finding potential applications in keratin hydrolysis and keratinase production.

Effect of glycosides of Cistanche on the expression of mitochondrial precursor protein and keratin type Ⅱ cytoskeletal 6A in a rat model of vascular dementia

Glycosides of Cistanche（GC）is a preparation used extensively for its neuroprotective effect against neurological diseases,but its mechanisms of action remains incompletely understood.Here,we established a bilateral common carotid artery occlusion model of vascular dementia in rats and injected the model rats with a suspension of GC（10 mg/kg/day,intraperitoneally）for 14 consecutive days.Immunohistochemistry showed that GC significantly reduced p-tau and amyloid beta（Aβ）immunoreactivity in the hippocampus of the model rats.Proteomic analysis demonstrated upregulation of mitochondrial precursor protein and downregulation of keratin type II cytoskeletal6A after GC treatment compared with model rats that had received saline.Western blot assay confirmed these findings.Our results suggest that the neuroprotective effect of GC in vascular dementia occurs via the promotion of neuronal cytoskeleton regeneration.

An ultrasonic-ionic liquid process for the efficient acid catalyzed hydrolysis of feather keratin

A novel efficient method for hydrolyzing feather keratin using an ultrasonic-ionic liquid coupling process has been developed, with reaction conditions optimized using response surface analysis of data obtained from single factor optimization studies. Ultrasonic irradiation(225 W power) of feathers in 8.4 mol·L^(-1) hydrochloric acid containing 1-butyl-3-methylimidazolium chloride as a cosolvent at 80 °C for 1.4 h, followed by heating at 110 °C for 8.3 h, resulted in hydrolysis of their keratin component in an excellent 83.1% yield. Compared with previous methods, this new method employs reduced amounts of hydrochloric acid, shorter reaction time, and affords amino acid hydrolysis products in higher yield.

Festosterone regulates keratin 33B expression in rat penis growth through androgen receptor signaling

Androgen therapy is the mainstay of treatment for the hypogonadotropic hypogonadal micropenis because it obviously enhances penis growth in prepubescent microphallic patients. However, the molecular mechanisms of androgen treatment leading to penis growth are still largely unknown. To clarify this well-known phenomenon, we successfully generated a castrated male Sprague Dawley rat model at puberty followed by testosterone administration. Interestingly, compared with the control group, testosterone treatment stimulated a dose-dependent increase of penis weight, length, and width in castrated rats accompanied with a dramatic recovery of the pathological changes of the penis. Mechanistically, testosterone administration substantially increased the expression of androgen receptor （AR） protein. Increased AR protein in the penis could subsequently initiate transcription of its target genes, including keratin 33B （Krt33b）. Importantly, we demonstrated that KRT33B is generally expressed in the rat penis and that most KRT33B expression is cytoplasmic. Furthermore, AR could directly modulate its expression by binding to a putative androgen response element sequence of the Krt33b promoter. Overall, this study reveals a novel mechanism facilitating penis growth after testosterone treatment in precastrated prepubescent animals, in which androgen enhances the expression of AR protein as well as its target genes, such as Krt33b.

Peri-implant keratinized gingiva augmentation using xenogeneic collagen matrix and platelet-rich fibrin:A case report

BACKGROUND Keratinized gingival insufficiency is a disease attributed to long-term tooth loss,can severely jeopardizes the long-term health of implants.A simple and effective augmentation surgery method should be urgently developed.CASE SUMMARY A healthy female patient,45-year-old,requested implant restoration of the her left mandibular first molar and second molar.Before considering a stage II,as suggested from the probing depth measurements,the widths of the mesial,medial,and distal buccal keratinized gingiva of second molar(tooth#37)were measured and found to be 0.5 mm,0.5 mm,and 0 mm,respectively.This suggested that the gingiva was insufficient to resist damage from bacterial and mechanical stimulation.Accordingly,modified apically repositioned flap(ARF)surgery combined with xenogeneic collagen matrix(XCM)and platelet-rich fibrin(PRF)was employed to increase the width of gingival tissue.After 1 mo of healing,the widths of mesial,medial,and distal buccal keratinized gingiva reached 4 mm,4 mm,and 3 mm,respectively,and the thickness of the augmented mucosa was 4.5 mm.Subsequently,through the second-stage operation,the patient obtained an ideal soft tissue shape around the implant.CONCLUSION For cases with keratinized gingiva widths around implants less than 2mmthe soft tissue width and thickness could be increased by modified ARF surgery combined with XCM and PRF.Moreover,this surgery significantly alleviated patients’pain and ameliorated oral functional comfort.

Fabrication and Evaluation of Porous Keratin/chitosan(KCS) Scaffolds for Effectively Accelerating Wound Healing

Objective To develop a dressing with desired antibacterial activity, good water maintaining ability and mechanical properties for wound healing and skin regeneration. Methods The chitosan with different concentrations were added in keratin solution to form porous keratin/chitosan（KCS） scaffolds. The morphological characteristics, chemical composition, wettability, porosity, swelling ratio and degradation of the scaffolds were evaluated. The antibacterial activity was tested by using S. aureus and E. coli suspension for 2 h. And L929 fibroblast cells culture was used to evaluate the cytotoxicity of the KCS scaffolds. Results The adding of chitosan could increase the hydrophobicity, decrease porosity, swelling ratio and degradation rate of the KCS porous scaffolds. Mechanical properties of KCS scaffolds could be enhanced and well adjusted by chitosan. KCS scaffolds could obviously decrease bacteria number. The proliferation of fibroblast cells in porous KCS patch increased firstly and then decreased with the increase of chitosan concentration. It was appropriate to add 400 μg/m L chitosan to form porous KCS scaffold for achieving best cell attachment and proliferation compared with other samples. Conclusion The porous KCS scaffold may be used as implanted scaffold materials for promoting wound healing and skin regeneration.

Association of keratin 8/18 variants with non-alcoholic fatty liver disease and insulin resistance in Chinese patients: A case-control study

AIM To test the hypothesis that K8/K18 variants predispose humans to non-alcoholic fatty liver disease(NAFLD) progression and its metabolic phenotypes. METHODS We selected a total of 373 unrelated adult subjects from our Physical Examination Department, including 200 unrelated NAFLD patients and 173 controls of both genders and different ages. Diagnoses of NAFLD were established according to ultrasonic signs of fatty liver. All subjects were tested for population characteristics, lipid profile, liver tests, as well as glucose tests. Genomic DNA was obtained from peripheral blood with a DNeasy Tissue Kit. K8/K18 coding regions were analyzed, including 15 exons and exon-intron boundaries. RESULTS Among 200 NAFLD patients, 10(5%) heterozygouscarriers of keratin variants were identified. There were 5 amino-acid-altering heterozygous variants and 6 non-coding heterozygous variants. One novel aminoacid-altering heterozygous variant(K18 N193S) and three novel non-coding variants were observed(K8 IVS5-9A→G, K8 IVS6+19G→A, K18 T195T). A total of 9 patients had a single variant and 1 patient had compound variants(K18 N193S+K8 IVS3-15C→G). Only one R341 H variant was found in the control group(1 of 173, 0.58%). The frequency of keratin variants in NAFLD patients was significantly higher than that in the control group(5% vs 0.58%, P = 0.015). Notably, the keratin variants were significantly associated with insulin resistance(IR) in NAFLD patients(8.86% in NAFLD patients with IR vs 2.5% in NAFLD patients without IR, P = 0.043).CONCLUSION K8/K18 variants are overrepresented in Chinese NAFLD patients and might accelerate liver fat storage through IR.

The Biocompatibility of Wool Keratin

Keratin is the major structural fibrous protein providing outer covering such as hair,wool,feathers,etc.When being used as a kind of biomaterials,the biocompatibility of wool keratin is one of the most critical questions.By now,there has not been systemic study on the biocompatibility of keratin.Therefore,in this article we used the procedures of skin irritation,haemolysis and subcutaneous implantation according to ISO 10993 to study it.Moreover,the Fourier transform-infrared(FTIR) spectroscopy was utilized to analyse the impurity and structure modification of wool keratin film.The part of the animal tests showed that the wool keratin films prepared by authors were biocompatible.But the residual of sodium dodecyl sulfate(SDS) affected the results of other tests.Consequently,the wool keratin membrane is one kind of favourable and promising biomaterial for biomedical and histological utilization.The residual SDS used as an agent should be eliminated from the keratin solution or membrane completely if for biological usage.In conclusion,wool keratin,as a kind of natural protein,prospectively could be applied in biomedical materials and scaffolds of tissue engineering.

Pleomorphic Adenoma with Exuberant Squamous Metaplasia and Keratin Cysts Mimicking Squamous Cell Carcinoma in Minor Salivary Gland

Salivary gland tumors, the second most common neoplasm of the mouth after squamous cell carcinoma, account for a significant proportion of tumors of the oral and perioral regions. An unusual case of adenoma presented as a solitary intraoral palatine mass in a 32-year-old woman is reported here. The tumor was interpreted as an unusual pleomorphic adenoma because of the presence of exuberant squamous metaplasia, clinically mimicking squamous cell carcinoma. Moreover, the presence of cystic structures filled with keratinized material was also salient feature. Pleomorphic adenomas may occasionally display focal squamous metaplastic changes;when extensive, it presents the potential for misinterpretation of the histology as indicative of well-differentiated squamous cell carcinoma.

The Study on the Expression of Keratin Proteins in Pterygial Epithelium

Purpose:To evaluate the expression patterns of several keratin proteins in pteygial epithelium.Methods:The proteins of K8,K10,K14,K16andAE3 in the epithelium of 8pterygia and 4normal conjunctiva were detected using immunofluorescent staining.Results:In pterygial epithelium,the staining patterns of 5keratin proteins were different with normal conjunctiva.No staining of K8,K10and K16was detected in normal conjunctival epithelium,but in pterygial epithelium,K8and K16were present in full thickness and K10was present in superficial cells.In normal conjunctiva.only base layer of epithelium showed positive staining with antibody of K14and AE3proteins,but they were present in the full thickness in pterygial epithelium and the antibody of AE3 protein stained the superficial cells heavily comparing with base cells.Conclusion:The Abnormal expression of keratin proteins(K8,K10,K14,K16andAE3)in pterygia indicates that the epithelial cells in pterygia are in the state of hyperproliferation and keratinization,which may,be the cause of the abnormal tear function in patients with pterygia,Eye Science2000;16:48-52.