Exploratory Analyses of Circulating Neoplastic-Immune Hybrid Cells as Prognostic Biomarkers in Advanced Intrahepatic Cholangiocarcinoma.

Existing clinical biomarkers do not reliably predict treatment response or disease progression in patients with advanced intrahepatic cholangiocarcinoma (ICC). Circulating neoplastic-immune hybrid cells (CHCs) have great promise as a blood-based biomarker for patients with advanced ICC. Peripheral blood specimens were longitudinally collected from patients with advanced ICC enrolled in the HELIX-1 phase II clinical trial (NCT04251715). CHCs were identified by co-expression of pan-cytokeratin (CK) and CD45, and levels were correlated to patient clinical disease course. Unsupervised machine learning was then performed to extract their morphological features to compare them across disease courses. Five patients were included in this study, with a median of nine specimens collected per patient. A median of 13.5 CHCs per 50,000 peripheral blood mononuclear cells were identified at baseline, and levels decreased to zero following the initiation of treatment in all patients. Counts remained undetectable in three patients who demonstrated end-of-trial clinical treatment response and conversely increased in two patients with evidence of therapeutic resistance. In the post-trial surveillance period, interval counts increased prior to or at the time of clinical progression in three patients and remain undetectable in one patient with continued long-term disease stability. Using our machine learning platform, treatment-resistant CHCs exhibited upregulation of CK and downregulation of CD45 relative to treatment-responsive CHCs. CHCs represent a promising blood-based biomarker to supplement traditional radiographic and biochemical measures.

New Bacillus paralicheniformis strain with high proteolytic and keratinolytic activity.

Bacillus paralicheniformis T7, which exhibits high proteolytic and keratinolytic activities, was isolated from soil in Kazakhstan. Its secreted proteases were thermostable and alkaline, demonstrating maximum activity at 70 °C and pH 9.0. The proteases and keratinases of this strain were sensitive to Ni2+, Co2+, Mn2+, and Cd2+, with Cu2+, Co2+ and Cd2+ negatively affecting keratinolytic activity, and Fe3+ ions have a strong inhibitory effect on proteolytic and keratinolytic activity. Seven proteases were identified in the enzymatic extract of B. paralicheniformis T7: four from the serine peptidase family and three from the metallopeptidase family. The proteases hydrolyzed 1 mg of casein, hemoglobin, gelatin, ovalbumin, bovine serum albumin, or keratin within 15 s to 30 min. The high keratinolytic activity of this strain was confirmed through the degradation of chicken feathers, horns, hooves, wool, and cattle hide. Chicken feathers were hydrolyzed in 4 days, and the degrees of hydrolysis for cattle hide, wool, hoof, and horn after 7 days of cultivation were 97.2, 34.5, 29.6, and 3.6%, respectively. During submerged fermentation with feather medium in a laboratory bioreactor, the strain secreted enzymes with 249.20 ± 7.88 U/mL protease activity after 24 h. Thus, B. paralicheniformis T7 can be used to produce proteolytic and keratinolytic enzymes for application in processing proteinaceous raw materials and keratinous animal waste.

A multicentre clinical trial evaluating the outcomes of two application regimens of a unique keratin-based graft in the treatment of Wagner grade one non-healing diabetic foot ulcers.

Diabetic foot complications that lead to lower extremity amputations pose a significant challenge to the entire global health system. In this multicentre clinical trial, 26 patients with chronic Wagner one diabetic foot ulcers (DFUs) were treated with a unique human keratin matrix graft applied either weekly or bi-weekly, in addition to standard of care. The hypothesis was that bi-weekly application would be similar to weekly application. The primary endpoint was complete wound closure by 12 weeks, and secondary endpoints included healing time, percent area reduction and weekly changes in peripheral neuropathy, pain and quality of life. In the intent-to-treat population, 77% (10/13) of DFUs treated with bi-weekly application healed compared with 69% (9/13) treated with weekly application. The mean time to heal within 12 weeks in the bi-weekly group was 61 days and in the weekly group was 54 days. The mean percent area reduction at 12 weeks was 94.7% in the bi-weekly group compared with 84.8% in the weekly group. The number of grafts used in the bi-weekly group was 3.9 compared with 6.2 in the weekly group. The results of this trial confirm our hypothesis that whether bi-weekly or weekly application of the unique keratin matrix graft is used to treat nonhealing indolent DFUs, there is a high rate of complete healing. Based on these results, future studies should be conducted that further investigate the use of this novel human keratin matrix graft for the treatment of chronic DFUs.

Characterization of a Bacterial Keratinolytic Protease for Effective Degradation of Chicken Feather Waste into Feather Protein Hydrolysates.

BACKGROUND: Chicken feathers contribute to large quantities of keratinaceous wastes that pose serious environmental problems and must be catered to properly. Chicken feathers are also a potential source of vital proteins, peptides, and amino acids, which could be used as low-cost animal feeds. Therefore, there has been increasing interest in keratinase-producing microbes for reprocessing and using keratinous biomaterials. METHODS: Among the five isolated keratinolytic microorganisms, one microbe, Bacillus XT 01, produced a significant amount of enzyme activity, which was partially characterized. The potential of this protease-producing microbe was investigated for converting feather keratin waste to valuable protein hydrolysate. RESULTS: Maximum keratinase production was observed after 5 days of incubating Bacillus XT 01 at an optimum temperature of 45 °C and pH 8.5. Sodium Dodecyl Sulphate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and zymogram of ammonium sulfate precipitated culture supernatant showed the presence of several proteolytic enzymes with molecular weights between 30 and 60 kDa. The Bacillus strain caused almost complete feather degradation (98%) after 7 days of incubation at 45 °C in a shake culture medium. Antioxidant and reducing activities of the feather protein hydrolysate (FPH) elevated with increased cultivation time. Investigation of the effect of feather protein hydrolysate on plants indicated improved plant growth regarding the agronomic parameters, such as plant height, number of trifoliate leaves, number of pods, pod length, number of seeds per pod, and root length, which increased by 30.84%, 49.32%, 70.90%, 53.27%, 60.03%, and 54.71%, respectively. CONCLUSIONS: The prospective of Bacillus XT 01 for degrading feather waste keratin to highly valued hydrolyzed feather protein offers effectiveness in the poultry industry and ultimately decreases environmental pollution hazards.

Mutation of KAP, which encodes a keratin-associated protein, affects grain size and yield production in rice.

Grain size and shape are critical agronomic traits that directly impact rice grain yield. Identifying genes that control these traits can provide new strategies for yield improvement. In this study, we characterized a rice mutant, reduced grain length (rgl), which exhibited decreased grain length due to reduced cell proliferation. Map-based cloning identified a base deletion in OsRGL2, a gene encoding a keratin-associated protein (KAP), as the cause of the mutant phenotype. CRISPR-Cas9-generated OsRGL2 knockout mutants also displayed reduced grain length, confirming its role. OsRGL2 transcripts were detected in various tissues, with relative higher gene expression in young panicles, and OsRGL2 was localized to the plasma membrane. Overexpression of OsRGL2 increased grain size by promoting cell proliferation in the spikelet hull and significantly enhanced grain yield per plant. Importantly, OsRGL2 was found to interact with RGB1, indicating that OsRGL2 positively regulates grain size and yield through its interaction with RGB1. Additionally, OsRGL2 regulated the expression of cell cycle-related genes, further elucidating its role in grain development. These findings demonstrate that OsRGL2 is a positive regulator of grain size in rice, and manipulating its expression may offer a novel strategy for enhancing rice grain yield.

A long-term study of stable isotope ratios of fingernail keratin and amino acids in a mother-infant dyad.

OBJECTIVE: To evaluate the potential of compound-specific isotope analysis of amino acids (CSIA-AA) for investigating infant feeding practices, we conducted a long-term study that compared infant and maternal amino acid (AA) nitrogen isotope ratios. MATERIALS AND METHODS: Fingernail samples were collected from a single mother-infant dyad over 19 months postpartum. Carbon and nitrogen stable isotope ratios were measured in the bulk keratin of the fingernail samples. Selected samples were then hydrolyzed and derivatized for compound-specific nitrogen isotope analysis of keratin AAs. RESULTS: As in previous studies, infant bulk keratin nitrogen isotope values increased during exclusive breastfeeding and fell with the introduction of complementary foods and eventual cessation of breastfeeding. Infant trophic AAs had elevated nitrogen isotope values relative to the mother, while the source AAs were similar between the mother and infant. Proline and threonine appeared to track the presence of human milk in the infant's diet as the isotopic composition of these AAs remained offset from maternal isotope values until the cessation of breastfeeding. DISCUSSION: Although CSIA-AA is costly and labor intensive, it appears to hold potential for estimating the duration of breastfeeding, even after the introduction of complementary foods. Through the analysis of a full suite of AAs, it may also yield insights into infant physiology and AA synthesis.

Low temperature carbonization and synergistic flame retardancy of cotton fabric coated by phosphorus­nitrogen flame retardants.

To solve the problems of flammability and smoldering of cotton fabric, its flame-retardant finishing was executed with biomass wool keratin (WK) and cyclic phosphate ester (CPE) through the soaking and baking process. The synergistic mechanism of WK low-temperature melting and CPE catalytic dehydration prompted the formation of protective carbonization layer on cotton fabric surface, and this protective layer reduced its pyrolysis rate, inhibited the production of combustible materials and improved its flame retardancy. The results of synchronous thermal analysis indicate that the initial decomposition temperature of WK and CPE is lower than that of cotton fabric, and they precede the endothermic degradation before fabric main body. This effectively promotes the low-temperature carbonization of cotton fabric and inhibits its pyrolysis. The initial decomposition temperature of WK/CPE treated fabrics advances by 47.9 °C-97.8 °C, presenting significant low-temperature carbonization trend. Moreover, they form 3.0 %-20.0 % aromatic structural char before the pyrolysis of cotton cellulose due to the low-temperature dehydration and carbonization reactions. The damage length after vertical burning is only 4.0 cm for treated fabric with five layers, its after-flame and smoldering disappear, and its limiting oxygen index value increases to 28.7 %. This research provides an effective idea for the flammability and smoldering problems of cotton fabric.

Design, development and characterization of a chimeric protein with disulfide reductase and protease domain showing keratinase activity.

Keratin is one of the major components of solid waste, and the degradation products have extensive applications in various commercial industries. Due to the complexity of the structure of keratin, especially the disulfide bonds between keratin polypeptides, keratinolytic activity is efficient with a mixture of proteins with proteases, peptidases, and oxidoreductase activity. The present work aimed to create an engineered chimeric protein with a disulfide reductase domain and a protease domain connected with a flexible linker. The structure, stability, and substrate interaction were analyzed using the protein modeling tools and codon-optimized synthetic gene cloned, expressed, and purified using Ni2+-NTA chromatography. The keratinolytic activity of the protein was at its maximum at 70 °C. The suitable pH for the enzyme activity was pH 8. While Ni2+, Mg2+, and Na+ inhibited the keratinolytic activity, Cu2+, Ca2+, and Mn2+ enhanced it significantly. Biochemical characterization of the protease domain indicated significant keratinolytic activity at 70 °C at pH 10.0 but was less efficient than the chimeric protein. Experiments using feathers as the substrate showed a clear degradation pattern in the SEM analysis. The samples collected from the degradation experiments indicated the release of proteins (2-fold) and amino acids (8.4-fold) in a time-dependent manner. Thus, the protease with an added disulfide reductase domain showed excellent keratin degradation activity and has the potential to be utilized in the commercial industries.

Effects of KRTAP20-1 Gene Variation on Wool Traits in Chinese Tan Sheep.

Chinese Tan sheep lambs are recognised for having tight 'spring-like' curly wool when young, but this phenotype disappears with age. This wool consists of shorter, fine wool fibres (which are usually unmedullated) and heterotypic hair fibres (which are frequently medullated), which are referred to as 'halo hair'. Both the wool and hair fibres consist of α-keratin proteins embedded in a keratin-associated protein (KAP) matrix. Of these KAPs, the KAP20-1 gene (designated KRTAP20-1) and its effect on four fibre traits (mean fibre curvature, mean fibre diameter, fibre diameter standard deviation, and coefficient of variation of fibre diameter) of Tan lambs was studied. Seven previously identified KRTAP20-1 variants (A, B, D, E, F, G, and H) of KRTAP20-1 were revealed, but the previously identified variant C was not present. Of the seven variants detected, only two (A and G) were common and present at frequencies greater than 5%, and the effect of these on the fibre traits of the finer wool fibres was assessed. It was found that variant G was associated with an increased mean fibre curvature in these wool fibres. This suggests that KRTAP20-1 might possibly be expressed differentially in the two fibre types, which may be of future value in breeding.

Changes of epidermal proteins immunolocalization in the corneous layer from embryonic to definitive avian beak.

The process of keratinization and cornification in the developing beak has been studied through immunofluorescence and immunogold electron microscopy in chick and zebrafinch embryos. After the curved beak anlagen appears at the tip of the maxillar bone, 5-8 layers of embryonic epidermis are generated from the basal layer of the epidermis. These cells are weakly immunoabeled for IFKs (Intermediate Filament Keratins) and more intensely for scaffoldin, a protein of the EDC (Epidermal Differentiation Complex) involved in the soft keratinization of the embryonic epidermis. Immunolabeling for CBPs (Corneous Beta Proteins) is visible in the transitional embryonic layers that are temporarily generated between the embryonic and definitive beak epidermis. The electron microscope reveals that intermediate layers contain immunolabeled periderm granules for scaffoldin mixed with bundles of corneous material immunolabeled for CBPs. Intense CBPs labeling occurs in the compacting corneous bundles of beta-keratinocytes in the definitive beak while scaffolding labeling disappears. The embryonic epidermis is sloughed before hatching. Sox (Sulfhydryl Oxidase) immunolabeling reveals that the enzyme is almost absent in embryonic layers but is present in transitional and definitive beta-keratinocytes. This indicates the formation of cross-linked disulfide bonds in the definitive corneous layer of the beak. Some calcium precipitation, suggested from von Kossa staining, occurs in the corneous layers only on the 18th day of development in the chick, in preparation for hatching.

The molecular mechanisms underlying optical isomer arbutin permeating the skin: The molecular interaction between arbutin and skin components.

Arbutin, a typical optical isomer, has garnered widespread acclaim in the whitening cosmetics for its favorable efficacy and safety. However, the molecular mechanisms underlying α-arbutin and ß-arbutin permeating across the skin have not elucidated clearly yet. Herein we aimed to unveil how α-arbutin and ß-arbutin interacted with keratin or SC lipids, further demonstrating their relationship with their drug permeability. We found that α-arbutin displayed significantly higher drug accumulation into the porcine skin than ß-arbutin within 24 h through in vitro permeation test. Moreover, α-arbutin predominantly induced the alternations of secondary structure of amide II during the drug permeation, which was favorable for α-arbutin permeation. On the contrary, ß-arbutin exhibited an observable effect on the stretching vibration of SC lipids, possessing a significantly stronger mixing energy, binding energy and compatibility with ceramide (Cer) than that of α-arbutin, which ultimately restricted its permeation. Interestingly, free fatty acids and ceramides of the SC lipids specifically utilized its oxygen atom of carboxyl group to dock the arbutin molecules, enhancing their affinity with ß-arbutin, as confirmed by molecular simulation and 13Carbon Nuclear Magnetic Resonance. Nevertheless, a favorable compatibility between α-arbutin and keratin was observed. It was emphasized that the distinct spatial configuration and opposite optical rotation of arbutin was the leading factor impacting the intermolecular force between arbutin and the SC, and resulted in a diverse drug permeation. In cellular and in vivo skin pharmacokinetic studies, α-arbutin also possessed a higher cellular uptake and topical bioavailability than ß-arbutin. This study revealed the transdermal permeation mechanisms of optical isomer arbutin at the molecular levels, providing methodological reference for the investigations of permeation behaviors of other isomers with similar spatial configuration.

Morphofunctional Characteristics of the Vessels of the Subareolar Lymphatic Plexus in Breast Cancer with Metastasis to the Lymph Nodes of the Axillary Lymphatic Collector.

The subareolar Sappey's plexus was studied using color lymphography and immunohistochemical methods with a panel of antibodies to podoplanin, smooth muscle actin, low molecular weight cytokeratin AE1/AE3, and GATA3 on archival material obtained during radical mastectomies and sectoral resections with lymph node dissection from 86 patients diagnosed with non-special type breast cancer. At the macro- and microscopic levels, the connection between the subareolar lymphatic plexus and the lymphatic system of the breast parenchyma has been demonstrated. In triple negative breast cancer with metastases to the axillary lymph nodes, the involvement of subareolar lymphatic plexus into lymphogenous metastasis to the lymph nodes of the axillary lymphatic collector was shown.

Development of a Combined Protein and Dye Extraction Approach for the Analysis of Keratin-Based Textiles.

Archaeological textiles represent precious remains from ancient culture; this is because of the historical and cultural importance of the information that can be obtained by such relics. However, the extremely complicated state of preservation of these textiles, which can be charred, partially or totally mineralized, with heavy soil or biological contamination, requires highly specialized and sensitive analytical tools to perform a comprehensive study. Starting from these considerations, the paper presents a combined workflow that provides the extraction of dyes and keratins and keratin-associated proteins in a single step, minimizing sampling while maximizing the amount of information gained. In the first phase, different approaches were tested and two different protocols were found suitable for the purpose of the unique workflow for dyes/keratin-proteins: a slightly modified urea protocol and a recently proposed new TCEP/CAA procedure. In the second step, after the extraction, different methods of cleanup and workflow for proteins and dyes were investigated to develop protocols that did not result in a loss of aliquots of the analytes of interest and to maximize the recovery of both components from the extracting solution. These protocols investigated the application of two types of paramagnetic beads, unmodified and carboxylate-coated hydrophilic magnetic beads, and dialysis and stage-tip protocols. The newly designed protocols have been applied to cochineal, weld, orchil, kermes, and indigo keratin-based dyed samples to evaluate the effectiveness of the protocols on several dye sources. These protocols, based on a single extraction step, show the possibility of investigating dyes and keratins from a unique sample of 1 mg or lesser, with respect to the thresholds of sensitivity and accuracy required in the study of textile artifacts of historical and artistic values.

Expression Patterns of Cytokeratins (CK7, CK20, CK19, CK AE1/AE3) in Atypical Endometrial Hyperplasia Coexisting with Endometrial Cancer.

Few studies have evaluated cytokeratin's (CK) staining patterns in atypical endometrial hyperplasia (AEH) coexisting with early-stage endometrial cancer (EC). We aimed to assess the staining patterns of selected CKs (CK7, CK19, CK20, CK AE1/AE3) in 74 patients with coexisting AEH and EC by independently analyzing both morphological variables. Specimens were collected from women with AEH and EC who underwent surgical interventions between 2012 and 2019 at the Department of Obstetrics and Gynecology of Vilnius University Hospital "Santaros Klinikos" in Vilnius, Lithuania. Immunostaining was also qualitatively classified as being heterogeneous or intense. The results revealed heterogeneous CK7 expression in all AEH cases and intense staining in 95.95% cases of AEH. The heterogeneous expression of CK7 was detected in all EC specimens. Intense CK7 expression was observed in 95.09% cases of EC G1 and in all G2 ECs. Heterogenous CK19 expression was present in all AEH specimens with intense staining in 92.42% of cases. Heterogeneous CK19 expression was observed in all EC samples with intense expression in 86.27% cases of EC G1 and 100% cases of EC G2. Interestingly, a significant relationship was found when comparing the heterogeneous expression of CK19 between AEH and well-differentiated EC. A significant difference was reported in the intense expression of CK AE1/AE3 (p = 0.031; p = 0.029) between AEH and G2 ECs and in the intense expression of CK AE1/AE3 between G1 and G2 ECs. CK20 staining was not a characteristic feature for AEH and early-stage EC. CK staining is present either in AEH or in early-stage endometrioid-subtype EC in different manners. Heterogeneous CK19 expression was significantly more common in AEH than in EC. CK20 expression was not associated with either AEH nor early-stage EC. An intense expression of CK AE1/AE3 was mainly present in moderately differentiated ECs, whereas the intense reactivity of AE1/AE3 showed a significant difference in well to moderately differentiated uterine tumors. The clinical implication of CK staining may aid in the more accurate diagnosis of AEH and early-stage EC as well as detect micrometastases leading to better oncological outcomes.

Is Raman Spectroscopy of Fingernails a Promising Tool for Diagnosing Systemic and Dermatological Diseases in Adult and Pediatric Populations?

Background: Raman spectroscopy is a well-known tool used in criminology, molecular biology, and histology. It is also applied to diagnose bone mineral disorders by taking advantage of the similarity of the structure of keratin and bone collagen. Raman spectroscopy can also be used in dermatology and diabetology. The purpose of the present review is to critically evaluate the available research about the use of Raman spectroscopy in the mentioned areas of medicine. Methodology: PubMed was searched for peer-reviewed articles on the subject of use of Raman spectroscopy in bone mineral disorders, dermatology, and diabetes mellitus. Results: Nail keratin and bone collagen are related structural proteins that require disulfide bond for structural stability. Therefore, Raman spectroscopy of keratin may have potential as a diagnostic tool for screening bone quality and distinguishing patients at risk of fracture for reasons different from low bone mineral density (BMD) in the adult women population. Raman spectroscopy can also investigate the changes in keratin's structure in nails affected by onychomycosis and distinguish between healthy and onychomycosis nail samples. It could also reduce the need for nail biopsy by distinguishing between dermatophytic and non-dermatophytic agents of onychomycosis. Additionally, Raman spectroscopy could expedite the diagnostic process in psoriasis (by assessing the secondary structure of keratin) and in diabetes mellitus (by examining the protein glycation level). Conclusions: In adult populations, Raman spectroscopy is a promising and safe method for assessing the structure of fingernails. However, data are scarce in the pediatric population; therefore, more studies are required in children.

Construction of artificial microbial consortia for efficient degradation of chicken feathers and optimization of degradation conditions.

Microbes within a consortium exhibit a synergistic interaction, enhancing their collective capacity to perform functions more effectively than a single species, especially in the degradation of keratin-rich substrates. To achieve a more stable and efficient breakdown of chicken feathers, a comprehensive screening of over 9,000 microbial strains was undertaken. This meticulous selection process identified strains with the capability to degrade keratin effectively. Subsequently, antagonistic tests were conducted to isolate strains of fungi and bacteria that were non-antagonistic, which were then used to form the artificial microbial consortia. The optimal fermentation conditions for the keratinophilic microbial consortia were determined through the optimization of response surface methodology. The results revealed that 11 microbial strains-comprising of 4 fungi and 7 bacteria-were particularly proficient in degrading chicken feathers. The artificially constructed microbial consortia (AMC) comprised two bacterial strains and one fungal strain. The optimal conditions for feathers degradation were identified as a 10 g/L concentration of chicken feathers, a 2.6% microbial inoculation volume and a fermentation fluid pH of 9. Under these conditions, the degradation rate for chicken feathers reached a significant 74.02%, representing an 11.45% increase over the pre-optimization rate. The AMC developed in this study demonstrates the potential for efficient and economical process of livestock and poultry feathers. It provides innovative insights and a theoretical foundation for tackling the challenging degradation of keratin-rich materials. Furthermore, this research lays the groundwork for the separation and purification of keratins, as well as the development of novel proteases, which could have profound implications for a range of applications.

Inopinatus corneliae sp. nov. gen. nov. isolated from human skin: A newly discovered keratinophilic hyphomycete, order Onygenales.

BACKGROUND: Fungi clinically relevant to human skin comprise prevalent commensals and well-known pathogens. Only rarely human skin harbours fungi that evade identification. OBJECTIVE: To characterise an enigmatic specimen isolated from a skin lesion. METHODS: A comprehensive clinical and mycological workup including conventional methods for phenotypic characterisation and sequencing based on internal transcribed spacer (ITS) and large subunit (LSU) regions to infer a phylogenetic tree. RESULTS: Cultures on common solid media were macroscopically inconspicuous initially until mycelial tufts developed on the surface, notably on potato dextrose agar. Polymorphous chlamydospores were detected but no aleurospores and ascomata. At 26°C, the isolate grew on standard agars, plant materials and garden soil and utilised peptone, keratins, lipids, inulin, erythrocytes and cellulose. It also grew at 5°C and at 37°C. Nucleotide sequences of its ITS region showed 93% similarity to sequences of different Malbranchea species. The closest matches among LSU rRNA sequences were obtained with the genera Amauroascus, Arthroderma, Auxarthronopsis and Malbranchea (93%-95%). A combined phylogenetic analysis placed the fungus in a sister clade to Neogymnomycetaceae, classified as incertae sedis in Onygenales, on a large distance to either Diploospora rosea or 'Amauroascus' aureus. CONCLUSIONS: The genus Inopinatus gen. nov. (MB854685) with the species Inopinatus corneliae sp. nov. (MB854687) is introduced to accommodate our isolate (holotype: DSM 116806; isotypes: CBS 151104, IHEM 29063). Probably Inopinatus corneliae is a geophilic species that, although potentially harmful, was no relevant pathogen in our case. Its ecology, epidemiology and pathogenicity need to be further clarified.

[Isolated intraductal carcinoma of the prostate: a clinicopathological and molecular analysis].

Objective: To study the clinicopathological features, immunohistochemical phenotypes, molecular changes, differential diagnosis and prognosis of isolated intraductal carcinoma of the prostate (iIDC-P). Methods: Three iIDC-P cases were collected retrospectively from 2016 to 2022 at Ningbo Clinical Pathology Diagnosis Center, Ningbo, China. The clinicopathologic features and immunophenotypic profiles were studied using light microscopy and immunohistochemistry. A targeted next-generation sequencing panel was used to analyze cancer-associated mutations. Follow-up and literature review were also performed. Results: The patients' ages were 61, 67 and 77 years, and their preoperative prostate specific antigen (PSA) levels were 7.99, 7.99 and 4.86 µg/L, respectively. Case 1 and 2 were diagnosed on needle biopsy and radical prostatectomy (RP) specimens, and case 3 was diagnosed on a specimen of transurethral resection of the prostate (TURP). The RP specimen was entirely submitted for histologic examination. In the case 1, iIDC-P was found in one tissue core (involving two ducts) in the biopsy specimen, and in 6 sections (diameter, 0.3-1.1 cm) from the radical prostatectomy specimen, and one section had separate foci of low-grade acinar adenocarcinoma (diameter, 0.05 cm). In the case 2, 6 tissue sections from the biopsy specimens showed iIDC-P, and 13 sections from RP specimen showed iIDC-P (diameter, 0.5-1.6 cm), and the other 3 sections had separate low grade acinar adenocarcinoma (diameter, 0.6 cm). In the case 3, 5 tissue blocks from the TURP specimen showed iIDC-P. The case 1 and 2 showed solid architecture with expansile proliferation of neoplastic cells in native ducts and acini. The case 3 showed dense or loose cribriform pattern, with marked cytological atypia, and frequent mitotic figures. Comedonecrosis was found in solid or dense cribriform glands in the case 2. Immunohistochemically, surrounding basal cells were highlighted using high-molecular-weight cytokeratin (34ßE12 and CK5/6) and p63, while P504s was positive in the tumor cells. The tumor cells were also positive for AR and prostate markers (NKX3.1, PSA and PSAP), and negative for GATA3. The iIDC-P and acinar adenocarcinoma both showed weak PTEN expression and no ERG (nuclear) expression. In case 2 and 3, targeted sequencing revealed activated oncogenic driver mutations in MAPK and PI3K pathway genes (KRAS, MTOR and PTEN). In addition, pathogenic mutation in TP53 and FOXA1 mutation were found in the case 2 and 3, respectively. No case demonstrated TMPRSS2::ERG translocation. All cases were microsatellite stable and had lower tumor mutation burdens (range, 2.1-3.1 muts/Mb). The patients showed no biochemical recurrence or metastasis after follow-up of 16-91 months. Conclusions: iIDC-P is a special type of intraductal carcinoma of the prostate and differs from intraductal carcinoma within high-grade prostate cancer. iIDC-P has unique molecular characteristics and may represent as a molecularly unique in situ tumor of prostate cancer.

Gingival open-wound to increase keratinized tissue in implant rehabilitation-A technical note.

Keratinized tissue augmentation around implants guarantees long-term success and maintenance of implant rehabilitations. Free gingival grafting is often described as the gold standard, especially when dealing with limited residual keratinized tissue height. Traditionally, an epithelio-conjunctive graft is harvested, either on the palate or the tuberosity, to reconstruct the missing keratinized soft tissues. This article introduces an innovative approach to increase keratinized tissue around implants, benefiting from second-intention gingival healing. This original surgical approach is interesting because it does not involve autogenous grafting or biomaterials. Its main goals are to enhance predictability while reducing the numerous per and post-operative risks related to autogenous harvesting. The success of this technique depends on the observance of fundamental principles: protection against bacterial contamination (immunocompetence of the patient), phenotypic induction (preoperative presence of keratinized tissue), space maintenance, and stabilization of flaps.

Tandem LIM domain-containing proteins, LIMK1 and LMO1, directly bind to force-bearing keratin intermediate filaments.

The cytoskeleton of the cell is constantly exposed to physical forces that regulate cellular functions. Selected members of the LIM (Lin-11, Isl-1, and Mec-3) domain-containing protein family accumulate along force-bearing actin fibers, with evidence supporting that the LIM domain is solely responsible for this force-induced interaction. However, LIM domain's force-induced interactions are not limited to actin. LIMK1 and LMO1, both containing only two tandem LIM domains, are recruited to force-bearing keratin fibers in epithelial cells. This unique recruitment is mediated by their LIM domains and regulated by the sequences outside the LIM domains. Based on in vitro reconstitution of this interaction, LIMK1 and LMO1 directly interact with stretched keratin 8/18 fibers. These results show that LIM domain's mechano-sensing abilities extend to the keratin cytoskeleton, highlighting the diverse role of LIM proteins in force-regulated signaling.