Efficacy and safety of combined UV-light corneal crosslinking and fine-needle diathermy to regress pathological murine corneal (lymph)angiogenesis in vivo.

PURPOSE: To compare safety and efficacy of isolated and combined UV-light corneal crosslinking (CXL) and fine-needle diathermy (FND) to regress pathological corneal vessels in vivo. METHODS: Mice with inflamed and pathologically vascularized corneas received CXL or FND as monotherapy or a combination of both treatments. Corneal pathological blood and lymphatic vessels, immune cells and the morphology of anterior segment structures were evaluated. RESULTS: All three approaches were able to regress blood and lymphatic vessels in mice. A comparative analysis of the three methods revealed that the FND monotherapy and the CXL + FND combination were significantly more effective than the CXL monotherapy, one and 2 weeks after therapy and especially in regressing lymphatic vessels. Furthermore, the combination therapy induced significantly less immune cell recruitment compared to the monotherapies. All three methods were safe to use in regards of corneal integrity. CONCLUSIONS: A combination of FND and CXL led to regression of pathological corneal lymphatic and blood vessels and reduced the infiltration of immune cells into inflamed murine corneas. This approach offers a new effective, safe and clinically usable strategy to treat eyes with mature pathological blood vessels and even more so for lymphatic vessels, for example prior to high-risk corneal transplantation.

ABCB5+ Limbal Epithelial Stem Cells Inhibit Developmental but Promote Inflammatory (Lymph) Angiogenesis While Preventing Corneal Inflammation.

The limbus, the vascularized junction between the cornea and conjunctiva, is thought to function as a barrier against corneal neovascularization. However, the exact mechanisms regulating this remain unknown. In this study, the limbal epithelial stem cell (LESC) marker ABCB5 was used to investigate the role of LESCs in corneal neovascularization. In an ABCB5KO model, a mild but significant increase of limbal lymphatic and blood vascular network complexity was observed in developing mice (4 weeks) but not in adult mice. Conversely, when using a cornea suture model, the WT animals exhibited a mild but significant increase in the number of lymphatic vessel sprouts compared to the ABCB5KO, suggesting a contextual anti-lymphangiogenic effect of ABCB5 on the limbal vasculature during development, but a pro-lymphangiogenic effect under inflammatory challenge in adulthood. In addition, conditioned media from ABCB5-positive cultured human limbal epithelial cells (ABCB5+) stimulated human blood and lymphatic endothelial cell proliferation and migration. Finally, a proteomic analysis demonstrated ABCB5+ cells have a pro(lymph)angiogenic as well as an anti-inflammatory profile. These data suggest a novel dual, context-dependent role of ABCB5+ LESCs, inhibiting developmental but promoting inflammatory (lymph)angiogenesis in adulthood and exerting anti-inflammatory effects. These findings are of high clinical relevance in relation to LESC therapy against blindness.

Regulatory T cells in rheumatoid arthritis: functions, development, regulation, and therapeutic potential.

Rheumatoid arthritis (RA) is an autoimmune disease that mainly affects the joints but also leads to systemic inflammation. Auto-reactivity and dysregulation of self-tolerance are thought to play a vital role in disease onset. In the pathogenesis of autoimmune diseases, disturbed immunosuppressive properties of regulatory T cells contribute to the dysregulation of immune homeostasis. In RA patients, the functions of Treg cells and their frequency are reduced. Therefore, focusing on the re-establishment of self-tolerance by increasing Treg cell frequencies and preventing a loss of function is a promising strategy for the treatment of RA. This approach could be especially beneficial for those patients who do not respond well to current therapies. In this review, we summarize and discuss the current knowledge about the function, differentiation and regulation of Treg cells in RA patients and in animal models of autoimmune arthritis. In addition, we highlight the therapeutic potential as well as the challenges of Treg cell targeting treatment strategies.

Effects of Femtosecond Laser-Assisted Cataract Surgery on Macular and Choroidal Thickness in Diabetic Patients.

INTRODUCTION: This study aimed to compare the short-term changes in retinal and choroid thickness in diabetic patients after femtosecond laser-assisted cataract surgery (FLACS) and phacoemulsification (PE) surgery. METHODS: A total of 47 eyes in the PE group and 44 eyes in the FLACS group were included. All patients underwent measurement of central macular thickness (CMT) and subfoveal choroidal thickness (SFCT) before and after surgery using optical coherence tomography (OCT). RESULTS: The effective phaco time (EPT) in the FLACS group was significantly reduced. The BCVA differed significantly between the two groups at 1 week and 1 month after surgery. The CMT in both groups increased at 1 week after the operation. It did not return to the preoperative level until month 12 in the PE group. In the FLACS group, the CMT began to decrease at month 3 and recovered to the preoperative level at month 12. The SFCT of the two groups increased at week 1; it began to decrease at month 6 in the PE group but did not recover to the preoperative level until month 12. The SFCT in the FLACS group recovered to preoperative levels at month 6. In the PE group, baseline CMT values predicted CMT change at week 1 and months 1, 3 and 12 after surgery. In the FLACS group, baseline CMT predicted CMT changes at week 1, month 1 and month 3. In the FLACS group, EPT predicted SFCT change at month 3. CONCLUSION: FLACS is safe and effective in patients with no fundus change or mild diabetic retinopathy. It has advantages in effectively reducing EPT, achieving good vision earlier and promoting faster recovery of the retinal and choroidal thickness. Preoperative CMT is a significant predictor of CMT changes in the early period after FLACS.

WRINKLED1 transcription factor orchestrates the regulation of carbon partitioning for C18:1 (oleic acid) accumulation in Siberian apricot kernel.

WRINKLED1 (WRI1), an APETALA2 (AP2)-type transcription factor, has been shown to be required for the regulation of carbon partitioning into fatty acid (FA) synthesis in plant seeds. To our knowledge, the regulatory network of WRI1 remains unknown in Prunus sibirica kernel (PSK), a novel woody biodiesel feedstock in China. In this study, based on the transcriptional data from developing oilseeds of multiple plant species, we identified 161 WRI1-coexpressed genes using weighted gene co-expression network analysis (WGCNA). The major portion of WRI1-coexpressed genes was characterized to be involved in carbon partitioning and FA biosynthesis. Additionally, we detected the temporal patterns for oil content and FA compositions in developing PSK from two different germplasms (AS-85 and AS-86). The major differences between the two germplasms are higher contents of oil and C18:1 in AS-85 than in AS-86 at a mature stage. Thus, AS-85 and AS-86 are desirable materials to explore the molecular and metabolic mechanisms of oil accumulation in Siberian apricot. Expression analysis in developing PSK of AS-85 and AS-86 indicated that the expression level of P. sibirica WRI1 (PsWRI1) was closely correlated to accumulative rate of oil. Also, the comparison of expression profiles in developing PSK of AS-85 and AS-86 displayed that the pPK, E1-α, E2, TAL, BC, MCMT, BS, SAD and FAD2 have a high correlation with PsWRI1. Transient expression showed that ProSAD- and ProBS-driving GUS expression showed no substantial difference between AS-85 and AS-86, while the expression level of ProPEPCK-AS-85 driving GUS was significantly higher than that of ProPEPCK-AS-86 driving GUS. Additionally, transient co-transformation with PsWRI1 revealed that ProSAD, ProPEPCK and ProBS activity could be specifically up-regulated by PsWRI1. This regulatory mechanism of PsWRI1 may create a steep concentration difference, thereby facilitating carbon flux into C18:1 accumulation in developing PSK. Overall, all our findings imply a versatile mechanism of WRI1 to optimize carbon allocation for oil accumulation, which can provide reference for researching the woody biodiesel plants.

Fruit characteristics, soluble sugar compositions and transcriptome analysis during the development of Citrus maxima "seedless", and identification of SUS and INV genes involved in sucrose degradation.

Citrus maxima "seedless" is originally from Malaysia, and now is widely cultivated in Hainan province, China. The essential features of this cultivar are thin skin, green epicarp and seedless at the ripening stage. Here, using C. maxima "seedless" as experimental material, we investigated the physical and inclusion indicators, and found the accumulation of storage compounds during 120-210 DAF leading to inconsistent increase between volume and weight. Component analysis of soluble sugar indicated that arabinose and xylose have a high content in early development of pummelo juice sacs (PJS), whereas fructose, glucose and sucrose show a significant increase during PJS maturation. To clarify a global overview of the gene expressing profiles, the PJSs from four periods (60, 120, 180 and 240 DAF) were selected for comparative transcriptome analysis. The resulting 8275 unigenes showed differential expression during PJS development. Also, the stability of 11 housekeeping genes were evaluated by geNorm method, resulting in a set of five genes (UBC, ACT, OR23, DWA2 and CYP21D) used as control for normalization of gene expression. Based on transcriptome data, 5 sucrose synthases (SUSs) and 10 invertases (INVs) were identified to be involved in sucrose degradation. Importantly, SUS4 may be responsible for arabinose and xylose biosynthesis to form the cell wall in early development, while SUS3 and VIN2 may be important in the accumulation of soluble hexose leading to cell expansion through an osmotic-independent pathway in late development. The information provides valuable metabolite and genetic resources in C. maxima "seedless", and is important for achieving high fruit yield and quality.

Physiological and transcriptomic analysis revealed the involvement of crucial factors in heat stress response of Rhododendron hainanense.

Molecular regulatory mechanism of heat stress response (HSR) in Ericaceae remains unknown. Here, we sought to identify HSR mechanisms in Rhododendron hainanense, a Ericaceae species, through a combination of physiological and transcriptomic studies. The levels of MDA, H2O2, Pro, SOD, CAT and APX in leaves of R. hainanense were analyzed to characterize a dramatic difference in varied temperature treatment. Also, three sequencing libraries, including one control and two heat stress (HS)-treated samples, were constructed for comparative transcriptomic analysis. By Illumina sequencing and Trinity strategy, 350 million clean reads (average length = 149 bp) was assembled into 183,486 unigenes. According to analysis of differential expression genes (DEGs), a total of 2658 DEGs were obtained. Moreover, a complex interaction network of 982 DEGs was established, of which master portions were comprised of 109 transcription factors (TFs). Importantly, integrated differential expression profiling, qRT-PCR and functional analysis, several TFs of R. hainanense (ABR1, IAA26, OBF1, LUX, SCL3, DIV, NAC29, NAC72 and TCP3) and their potential regulations for the crosstalk between hormonal signal and HSR were identified. These findings will contribute to our understanding of the regulatory mechanisms of HSR in R. hainanense, breeding cultivars with improved thermotolerance.

Identification of AUXIN RESPONSE FACTOR gene family from Prunus sibirica and its expression analysis during mesocarp and kernel development.

BACKGROUND: Auxin response factors (ARFs) in auxin signaling pathway are an important component that can regulate the transcription of auxin-responsive genes involved in almost all aspects of plant growth and development. To our knowledge, the comprehensive and systematic characterization of ARF genes has never been reported in Prunus sibirica, a novel woody biodiesel feedstock in China. RESULTS: In this study, we identified 14 PsARF genes with a perfect open reading frame (ORF) in P. sibirica by using its previous transcriptomic data. Conserved motif analysis showed that all identified PsARF proteins had typical DNA-binding and ARF domain, but 5 members (PsARF3, 8 10, 16 and 17) lacked the dimerization domain. Phylogenetic analysis of the ARF proteins generated from various plant species indicated that ARFs could be categorized into 4 major groups (Class I, II, III and IV), in which all identified ARFs from P. sibirica showed a closest relationship with those from P. mume. Comparison of the expression profiles of 14 PsARF genes in different developmental stages of Siberian apricot mesocarp (SAM) and kernel (SAK) reflected distinct temporal or spatial expression patterns for PsARF genes. Additionally, based on the expressed data from fruit and seed development of multiple plant species, we identified 1514 ARF-correlated genes using weighted gene co-expression network analysis (WGCNA). And the major portion of ARF-correlated gene was characterized to be involved in protein, nucleic acid and carbohydrate metabolic, transport and regulatory processes. CONCLUSIONS: In summary, we systematically and comprehensively analyzed the structure, expression pattern and co-expression network of ARF gene family in P. sibirica. All our findings provide theoretical foundation for the PsARF gene family and will pave the way for elucidating the precise role of PsARF genes in SAM and SAK development.