Allergen desensitization reduces the severity of relapsed alopecia areata in dust-mite allergic patients.

Atopy may be a facilitating factor in some alopecia areata (AA) patients with early disease onset and more severe/extensive AA. The underlying immune mechanisms are unknown, but allergen responses may support a pro-inflammatory environment that indirectly promotes AA. To investigate the long-term effect of allergen immunotherapy (AIT) against house dust mite (HDM) allergy on disease severity and prognosis for AA patients. An observational comparative effectiveness study was conducted on 69 AA patients with HDM allergy. 34 patients received conventional/traditional AA treatment (TrAA) plus AIT (AIT-TrAA), and 35 patients received TrAA alone. Serum total immunoglobulin E (tIgE), HDM specific IgE (sIgE), HDM specific IgG4 (sIgG4) and cytokines (IL-4, IL-5, IL-10, IL-12, IL-13, IL-33, IFNγ) were quantified in these patients, together with 58 non-allergic AA patients and 40 healthy controls. At the end of the 3-year desensitization course, the AIT-TrAA group presented with lower SALT scores than the TrAA group, especially in non-alopecia totalis/universalis (AT/U) patients and pre-adolescent AT/U patients (age ≤ 14). In patients with elevated tIgE levels before AIT, a decrease in tIgE was correlated to reduced extent of AA on completion of the AIT course. After desensitization, elevation of IL-5 and decrease of IL-33 were observed in HDM allergic-AA patients. Desensitization to HDM in allergic AA patients reduces the severity of relapse-related hair loss over the 3-year AIT treatment course, possibly via opposing Th2 dominance. This adjunctive treatment may help reduce disease severity and curtail the disease process in allergic patients with AA.

Simultaneously optimizing multiple properties of ß-glucosidase Bgl6 using combined (semi-)rational design strategies and investigation of the underlying mechanisms.

The performance of ß-glucosidase during cellulose saccharification is determined by thermostability, activity and glucose tolerance. However, conflicts between them make it challenging to simultaneously optimize three properties. In this work, such a case was reported using Bgl6-M3 as a starting point. Firstly, four thermostability-enhancing mutations were obtained using computer-aided engineering strategies (mutant M7). Secondly, substrate binding pocket of M7 was reshaped, generating two mutations that increased activity but decreased glucose tolerance (mutant M9). Then a key region lining active site cavity was redesigned, resulting in three mutations that boosted glucose tolerance and activity. Finally, mutant M12 with simultaneously improved thermostability (half-life of 20-fold), activity (kcat/Km of 5.6-fold) and glucose tolerance (ΔIC50 of 200 mM) was obtained. Mechanisms for property improvement were elucidated by structural analysis and molecular dynamics simulations. Overall, the strategies used here and new insights into the underlying mechanisms may provide guidance for multi-property engineering of other enzymes.

Sorption of organochlorine pesticides on polyethylene microplastics in soil suspension.

As a new type of environmental pollutant, microplastics (MPs) can adsorb residual organochlorine pesticides (OCPs) in the soil and pose a severe threat to the soil ecosystems. To understand the interaction between soil MPs and OCPs, the sorption of two kinds of OCPs, including hexachlorocyclohexanes (HCHs) and dichlorodiphenyltrichloroethanes (DDTs), on polyethylene (PE) microplastics in soil suspension was studied through sorption kinetics and isotherm models. The effects of solution/soil ratio and MPs diameter on sorption were examined. The kinetic experiment results show that the sorption equilibrium was 12 h， and the sorption process of OCPs on MPs can be well described by a pseudo-second-order model. The Freundlich model (R2 = 0.942-0.997) provides a better fit to the sorption isotherm data than the Langmuir model (R2 = 0.062-0.634), indicating that the sorption process takes place on the nonuniform surface of MPs. The MPs had a good sorption effect on OCPs when the solution/soil ratio was from 75:1 to 100:1. As the diameter of MPs increases, the sorption capacity decreases. These results provide support for further research on microplastic pollution in soil.

Nitric oxide alleviates lignification and softening of water bamboo (Zizania latifolia) shoots during postharvest storage.

Water bamboo shoots quickly deteriorate after harvest as a result of rapid lignification and softening. Nitric oxide (NO) has been used to extend the postharvest life of several other vegetables. Here, we examined the effect of NO on the storage of water bamboo shoots at 4℃ for 28 days. Without NO, fresh weight and firmness decreased quickly, while the cellulose and lignin contents increased sharply during storage. NO treatment delayed softening by maintaining the integrity of the cell wall and inhibiting the degradation of protopectin and the expressions of pectin methylesterase and polygalacturonase. NO treatment also delayed cellulose synthesis by increasing cellulase activity. NO treatment decreased the synthesis of lignin by inhibiting the activities of phenylalanine ammonia-lyase, cinnamyl alcohol dehydrogenase, laccase and peroxidase. These results indicate that NO treatment is effective at suppressing the softening and lignification of water bamboo shoots during postharvest storage.

Engineering of ß-Glucosidase Bgl15 with Simultaneously Enhanced Glucose Tolerance and Thermostability To Improve Its Performance in High-Solid Cellulose Hydrolysis.

In this study, a Petri-dish-based double-layer high-throughput screening method was established to improve glucose tolerance of ß-glucosidase Bgl15. Two beneficial mutations were identified, and the joint mutant 2R1 improved the half-maximal inhibitory concentration of glucose from 0.04 to 2.1 M. The crystal structure of 2R1 was subsequently determined at 2.7 Å. Structure analysis revealed that enhancement of glucose tolerance may be due to improved transglycosylation activity made possible by a hydrophobic binding site for glucose as an acceptor and more stringent control of a putative water channel. To further ameliorate the application potential of the enzyme, it was engineered to increase the half-life at 50 °C from 0.8 h (Bgl15) to 180 h (mutant 5R1). Furthermore, supplementation of 5R1 to the cellulase cocktail significantly improved glucose production from pretreated sugar cane bagasse by 38%. Consequently, this study provided an efficient approach to enhance glucose tolerance and generated a promising catalyst for cellulose saccharification.

The function of histone lysine methylation related SET domain group proteins in plants.

Histone methylation, which is mediated by the histone lysine (K) methyltransferases (HKMTases), is a mechanism associated with many pathways in eukaryotes. Most HKMTases have a conserved SET (Su(var) 3-9,E(z),Trithorax) domain, while the HKMTases with SET domains are called the SET domain group (SDG) proteins. In plants, only SDG proteins can work as HKMTases. In this review, we introduced the classification of SDG family proteins in plants and the structural characteristics of each subfamily, surmise the functions of SDG family members in plant growth and development processes, including pollen and female gametophyte development, flowering, plant morphology and the responses to stresses. This review will help researchers better understand the SDG proteins and histone methylation in plants and lay a basic foundation for further studies on SDG proteins.

Serum level of IL-4 predicts response to topical immunotherapy with diphenylcyclopropenone in alopecia areata.

BACKGROUND: This study investigated predictors of response to topical diphenylyclopropenone (DPCP) immunotherapy in patients with alopecia areata (AA). OBJECTIVE: To identify predictors of response, or resistance, to treatment for AA through clinical observations and serum tests. METHODS: Eighty four AA patients were treated with DPCP. Serum cytokine levels were measured in 33 AA patients pre- and post-treatment, and in 18 healthy controls, using ELISA assays. RESULTS: Of patients, 56.1% responded to DPCP with satisfactory hair regrowth; the response rate was negatively correlated with hair loss extent. Before DPCP treatment, higher serum IFN-γ and IL-12 cytokine levels were observed in AA patients compared to healthy controls. Non-responders to DPCP had significantly elevated serum IL-4 pre-treatment (3.07 fold higher) and lower IL-12 levels compared with responders. After DPCP treatment, non-responders had persistently high IL-4, increased IL-12, negligible decrease in IFN-γ and decreased IL-10. Post-treatment DPCP responders exhibited significantly decreased IFN-γ and IL-12, and increased IL-4 and IL-10. Development of adverse side-effects was significantly associated with higher pre-treatment serum IgE levels. LIMITATIONS: A small number of subjects were evaluated. CONCLUSIONS: Potentially, elevated pre-treatment serum levels of IL-4 and IL-12 can be used as unfavorable and favorable predictors of DPCP therapeutic effect, respectively. In addition, pre-treatment elevated serum total IgE may predict increased risk for severe adverse side-effects to DPCP application. Whether serum cytokine expression levels can be used as predictors of response to other forms of treatment is unknown, but it may warrant investigation in the development of personalized treatments for AA.

Improving Galactooligosaccharide Synthesis Efficiency of ß-Galactosidase Bgal1-3 by Reshaping the Active Site with an Intelligent Hydrophobic Amino Acid Scanning.

There are ongoing interests in improving the galactooligosaccharide (GOS) synthesis efficiency of ß-galactosidase by protein engineering. In this study, an intelligent double-hydrophobic amino acid scanning strategy was proposed and employed to target nine residues forming the glycon-binding site (-1 subsite) of ß-galactosidase Bgal1-3. Two mutants C510V and H512I with significantly improved GOS synthesis efficiency were obtained. When 40% (w/v) lactose was used as a substrate, Bgal1-3 reached a maximum GOS yield of 45.3% at 16 h, while the mutants reached higher yields in a much shorter time (59.1% at 10 h for C510V, 51.5% at 2 h for H512I). When skim milk was treated with these enzymes, more GOS was produced (19.9 g/L for C510V, 12.7 g/L for H512I) than that for Bgal1-3 (10.3 g/L) at a lactose conversion of 90%. These results validated hydrophobicity scanning as an efficient method to engineer ß-galactosidases into promising catalysts for the preparation of GOS and GOS-enriched milk.

Enhancing the Thermostability of Highly Active and Glucose-Tolerant ß-Glucosidase Ks5A7 by Directed Evolution for Good Performance of Three Properties.

A high-performance ß-glucosidase for efficient cellulose hydrolysis needs to excel in thermostability, catalytic efficiency, and resistance to glucose inhibition. However, it is challenging to achieve superb properties in all three aspects in a single enzyme. In this study, a hyperactive and glucose-tolerant ß-glucosidase Ks5A7 was employed as the starting point. Four rounds of random mutagenesis were then performed, giving rise to a thermostable mutant 4R1 with five amino acid substitutions. The half-life of 4R1 at 50 °C is 8640-fold that of Ks5A7 (144 h vs 1 min). Meanwhile, 4R1 had a higher specific activity (374.26 vs 243.18 units·mg-1) than the wild type with a similar glucose tolerance. When supplemented to Celluclast 1.5L, the mutant significantly enhanced the hydrolysis of pretreated sugar cane bagasse, improving the released glucose concentration by 44%. With excellent performance in thermostability, activity, and glucose tolerance, 4R1 will serve as an exceptional catalyst for industrial applications.

Tat-Independent Secretion of Polyethylene Terephthalate Hydrolase PETase in Bacillus subtilis 168 Mediated by Its Native Signal Peptide.

Widespread utilization of polyethylene terephthalate (PET) has caused critical environmental pollution. The enzymatic degradation of PET is a promising solution to this problem. In this study, PETase, which exhibits much higher PET-hydrolytic activity than other enzymes, was successfully secreted into extracellular milieu from Bacillus subtilis 168 under the direction of its native signal peptide (named SPPETase). SPPETase is predicted to be a twin-arginine signal peptide. Intriguingly, inactivation of twin-arginine translocation (Tat) complexes improved the secretion amount by 3.8-fold, indicating that PETase was exported via Tat-independent pathway. To the best of our knowledge, this is the first report on the improvement of Tat-independent secretion by inactivating Tat components of B. subtilis 168 in LB medium. Furthermore, PET film degradation assay showed that the secreted PETase was fully active. This study paves the first step to construct an efficient engineered strain for PET degradation.

Lesional infiltration of mast cells, Langerhans cells, T cells and local cytokine profiles in alopecia areata.

Alopecia areata (AA) is a chronic inflammatory disease mediated by an array of cells and cytokines. Immunohistochemistry (IHC) of histological sections with antibodies to mast cell tryptase, CD4, CD8, CD1a and semi-quantitative real-time PCR analysis of Th1- and Th2-type cytokines were performed in 55 patients to investigate the infiltration features of mast cells (MCs), T lymphocytes and Langerhans cells (LCs) in scalp lesions of patients with AA. In AA patients, increased MCs mainly infiltrated the peri-follicular and peri-vascular areas, and correlated positively with numbers of CD8(+) T lymphocytes in deep peri-follicular areas (P = 0.04), but negatively with CD4(+) T lymphocytes in deep peri-vascular areas (P = 0.031). In patients with active hair loss, LCs in epidermis, deep dermis and peri-vascular were elevated (Ps < 0.05). Infiltration of LCs in upper peri-vascular areas and CD8(+) T cell infiltration in deep peri-follicular areas were positively correlated (R = 0.618, P = 0.011), as well as LCs in deep peri-vascular areas with CD8(+) T cells in upper peri-follicular areas (R = 0.570, P = 0.017). In patients with active hair loss, Th1-type cytokine (IL-2, IL-8, TNF-α) mRNA expression in deep dermis were higher than in upper dermis (Ps < 0.05). However, in patients with non-active hair loss, Th2-type cytokine (IL-5, IL-10) mRNA expression in deep dermis was higher than that in the upper dermis (Ps < 0.05). Positive correlations were found existing between MCs and CD8(+) T cells, as well as between LCs and CD8(+) T cells. In conclusion, findings in this study allow us to propose a close relationship between mast cells and CD8(+) T cells, as well as between LCs and CD8(+) T cells in AA, as well as allergy may interfere with infiltrating T lymphocytes in AA lesional regions. Also, Th1-type cytokine are related to disease activity of alopecia areata, whereas Th2-type cytokines may be associated with persistence of AA.

Clinical features of primary cicatricial alopecia in Chinese patients.

BACKGROUND: There have been few reports on primary cicatricial alopecias (PCR) especially from Asia (PCA). AIMS: To study the clinical, pathological and dermoscopic characteristics of PCA among Chinese patients. METHODS: A retrospective analysis of the clinical data of 59 patients with PCA was conducted and the dermoscopic, pathological, treatment and prognosis characteristics analyzed. Fisher's Chi-square exact test, Kruskal-Wallis and Spearman rank correlation test were performed. RESULTS: The ratio of neutrophilic to lymphocytic cicatricial alopecias was about 1.3:1 in this group. The most frequent disorder was folliculitis decalvans. Follicular openings were absent on dermoscopy in all cases except alopecia mucinosa. Patulous follicular openings were characterisitc of alopecia mucinosa. After treatment, an increase in short vellus hairs was the earliest feature, while telangiectasia, epidermal scale, follicular hyperkeratosis, pustules and hair diameter diversity gradually decreased or even disappeared. Improvement in the areas of hair loss after treatment was seen more often in discoid lupus erythematosus, folliculitis decalvans and dissecting cellulitis than in patients with classic pseudopelade of Brocq. Nine patients (13.6%) relapsed after cessation of therapy. Female patients needed longer treatment times. Long duration, large areas of hair loss and shorter treatment courses were the major factors in relapses. CONCLUSIONS: Dermatoscopy provides a rapid, practical and useful aid for the diagnosis of PCA and also to assess disease activity. Patulous follicular openings are a specific dermoscopic sign of alopecia mucinosa. Lichen planopilaris is less common in China than in the West.