Improved chloroplast Pi allocation helps sustain electron transfer to enhance photosynthetic low-phosphorus tolerance of wheat.

Phosphorus (P) deficit limits high wheat (Triticum aestivum L.) yields. Breeding low-P-tolerant cultivars is vital for sustainable agriculture and food security, but the low-P adaptation mechanisms are largely not understood. Two wheat cultivars, ND2419 (low-P-tolerant) and ZM366 (low-P-sensitive) were used in this study. They were grown under hydroponic conditions with low-P (0.015 mM) or normal-P (1 mM). Low-P suppressed biomass accumulation and net photosynthetic rate (A) in both cultivars, whereas ND2419 was relatively less suppressed. Intercellular CO2 concentration did not decrease with the decline of stomatal conductance. Additionally, maximum electron transfer rate (Jmax) decreased sooner than maximum carboxylation rate (Vcmax). Results indicate that impeded electron transfer is directly responsible for decreased A. Under low-P, ND2419 exhibited greater PSII functionality (potential activity (Fv/Fo), maximum quantum efficiency (Fv/Fm), photochemical quenching (qL) and non-photochemical quenching (NPQ) required for electron transfer than ZM366, resulting more ATP for Rubisco activation. Furthermore, ND2419 maintained higher chloroplast Pi concentrations by enhancing chloroplast Pi allocation, compared with ZM366. Overall, the low-P-tolerant cultivar sustained electron transfer under low-P by enhancing chloroplast Pi allocation, allowing more ATP synthesis for Rubisco activation, ultimately presenting stronger photosynthesis capacities. The improved chloroplasts Pi allocation may provide new insights into improve low-P tolerance.

A Half-Sandwich Ruthenium(II) (N

Efficacy of clinical chemotherapeutic agents depends not only on direct cytostatic and cytotoxic effects but also involves in eliciting (re)activation of tumour immune effects. One way to provoke long-lasting antitumour immunity is coined as immunogenic cell death (ICD), exploiting the host immune system against tumour cells as a "second hit". Although metal-based antitumour complexes hold promise as potential chemotherapeutic agents, ruthenium (Ru)-based ICD inducers remain sparse. Herein, we report a half-sandwich complex Ru(II) bearing aryl-bis(imino) acenaphthene chelating ligand with ICD inducing properties for melanoma in vitro and in vivo. Complex Ru(II) displays strong anti-proliferative potency and potential cell migration inhibition against melanoma cell lines. Importantly, complex Ru(II) drives the multiple biochemical hallmarks of ICD in melanoma cells, i. e., the elevated expression of calreticulin (CRT), high mobility group box 1 (HMGB1), Hsp70 and secretion of ATP, followed by the decreased expression of phosphorylation of Stat3. In vivo the inhibition of tumour growth in prophylactic tumour vaccination model further confirms that mice with complex Ru(II)-treated dying cells lead to activate adaptive immune responses and anti-tumour immunity by the activation of ICD in melanoma cells. Mechanisms of action studies show that complex Ru(II)-induced ICD could be associated with mitochondrial damage, ER stress and impairment of metabolic status in melanoma cells. We believe that the half-sandwich complex Ru(II) as an ICD inducer in this work will help to design new half-sandwich Ru-based organometallic complexes with immunomodulatory response in melanoma treatments.

Nitrogen enhances the effect of pre-drought priming against post-anthesis drought stress by regulating starch and protein formation in wheat.

Drought stress is one of the most serious environmental stress factor constraining crop production across the globe. Among cereals, wheat grains are very sensitive to drought as a small degree of stress can affect the enzymatic system. This study aimed to investigate whether nitrogen and pre-anthesis drought priming could enhance the action of major regulatory enzymes involved in starch accumulation and protein synthesis in bread wheat (Triticum aestivum L.). For this purpose, cultivars YM-158 (medium gluten) and YM-22 (low gluten) were grown in rain-controlled conditions under two nitrogen levels, that is, N180 (N1) and N300 (N2). Drought priming was applied at the jointing stage and drought stress was applied 7 days after anthesis. Drought stress reduced starch content but enhanced protein content in grains. N2 and primed plants kept higher contents of nonstructural carbohydrates, fructans, and sucrose; with higher activity of sucrose-phosphate synthase in flag leaves. Furthermore, N2 and priming treatments showed higher sink ability to develop grains by showing higher sucrose-to-starch conversion activities of adenosine diphosphate-glucose pyrophosphorylase, uridine diphosphate glucose pyrophosphorylase, sucrose-synthase, soluble-starch synthase, starch branching enzyme, and granule-bound starch synthase as compared to N1 and non-primed treatments. The application of N2 and primed treatment showed a greater ability to maintain grain filling in both cultivars as compared to N1 and non-primed crops. Our study suggested that high nitrogen has the potential to enhance the effect of pre-drought priming to change source-sink relationships and grain yield of wheat under drought stress during the filling process.

Serum Expression of IL-33 and ST2 in Patients with Psoriasis Vulgaris.

BACKGROUND: Psoriasis vulgaris (PsV) is an immune-mediated skin disease of unknown mechanism. Interleukin 33 (IL-33) is a member of IL-1 cytokine family and suppression of tumorigenicity 2 (ST2) is the specific ligand of IL-33. It has been found that IL-33 and ST2 are increased in psoriatic lesions, but the expression levels in serum and their relationship to clinical features are still unclear. The aim of this study is to assess IL-33, ST2, IL-17 and IL-5 serum levels as well as serum concentration of blood glucose and blood lipids in PsV patients and their relationship with clinical characteristics. METHODS: Sixty-eight PsV samples and 60 healthy individuals were recruited. Serum levels of IL-33, ST2, IL-17 and IL-5 were measured by enzyme-linked immunosorbent assay and blood glucose and blood lipid were assayed by automatic biochemical analyzer. RESULTS: Serum levels of IL-33, ST2, IL-17 and IL-5 were increased significantly in PsV patients compared with controls (P<0.01). Cytokines were overexpressed in PsV patients during active stages compared with controls (P<0.05). Expression levels of IL-33, ST2 and IL-17 confirmed a significance in different severity groups of PsV patients (P<0.05). Serum concentration of triglyceride (TG) was also increased compared with controls (P=0.024). IL-33 levels were positively correlated with total cholesterol (TC) levels (r=0.319, P=0.008). CONCLUSION: IL-33/ST2 could generally reflect the activity and disease severity in PsV patients, which indicates that the IL-33/ST2 signaling pathway plays an important role in the pathogenesis of PsV.

A self-designed CpG ODN enhanced the anti-melanoma effect of pimozide.

Melanomas represent the deadliest form of skin cancers. Due to the intricacy of tumorigenesis, it is emergent to find effective therapies for melanomas. Researches have proved that pimozide inhibits the growth of melanoma, but the limited curing effect needs to be further improved. Nowadays, tumor immunotherapy has been widely recognized as the sole therapy that can eradicate cancers. Cytosine-phosphate-guanine oligodeoxynucleotide (CpG ODN), TLR9 receptor agonist, can significantly enhance anti-tumor immune responses. This study explored the therapeutic effect of pimozide combined with CpG ODN on melanoma-bearing mice. The results showed that pimozide combined with CpG ODN effectively inhibited the growth of melanoma and prolonged the survival of melanoma-bearing mice, inhibited the expression of MMP2 and p-Stat5, increased the infiltration of CD4+ and CD8+ T cells in tumor, raised the ratios of CD4+, CD8+ T cells and NK cells. These all indicated that the combination treatment improved the anti-tumor effect of pimozide on mice. The anti-tumor mechanism might be attributed to cell apoptosis induction, invasion inhibition, and immune regulation. A more effective combination treatment concerning with pimozide is being under investigation.

IL-33 contributes to disease severity in Psoriasis-like models of mouse.

Immune cells infiltrating the psoriatic skin secrete high amounts of pro-inflammatory cytokines IL-17, TNF-α, IL-21 and IL-36 resulting in chronic inflammation. However, the exact cellular and molecular mechanisms have not been fully understood. We report here elevation of IL-33 expression in psoriatic lesions. Studies in imiquimod (IMQ)-induced mice with psoriatic inflammation confirmed a critical role for IL-33 in driving the disease. IL-33 reduces the CD4+ and CD8+ cells, inhibits autophagy in IMQ-treated mouse skin, and promoted tyrosyl phosphorylation of STAT3. Thus, IL-33 appears to be a major risk factor for severity of psoriasis-like skin inflammation. Our findings may open new perspectives for understanding the mechanisms and developing a therapeutic strategy for psoriasis.

The enhanced antitumour response of pimozide combined with the IDO inhibitor L­MT in melanoma.

Melanoma is one of the most fatal and therapy-resistant types of cancer; therefore, identifying novel therapeutic candidates to improve patient survival is an ongoing effort. Previous studies have revealed that pimozide is not sufficient to treat melanoma; therefore, enhancing the treatment is necessary. Indoleamine 2, 3­dioxygenase (IDO) is an immunosuppressive, intracellular rate-limiting enzyme, which contributes to immune tolerance in various tumours, including melanoma, and inhibition of IDO may be considered a novel therapeutic strategy when combined with pimozide. The present study aimed to assess the antitumour activities of pimozide in vitro, and to investigate the effects of pimozide combined with L­methyl-tryptophan (L­MT) in vivo. For in vitro analyses, the B16 melanoma cell line was used. Cell cytotoxicity assay, cell viability assay, wound­healing assay and western blotting were conducted to analyse the effects of pimozide on B16 cells. Furthermore, B16 cell-bearing mice were established as the animal model. Haematoxylin and eosin staining, immunohistochemistry, terminal deoxynucleotidyl transferase dUTP nick end-labelling staining, western blotting and flow cytometry were performed to determine the effects of monotherapy and pimozide and L­MT cotreatment on melanoma. The results demonstrated that pimozide exhibited potent antitumour activity via the regulation of proliferation, apoptosis and migration. Furthermore, the antitumour effects of pimozide were enhanced when combined with L­MT, not only via regulation of proliferation, apoptosis and migration, but also via immune modulation. Notably, pimozide may regulate tumour immunity through inhibiting the activities of signal transducer and activator of transcription (Stat)3 and Stat5. In conclusion, the present study proposed the use of pimozide in combination with the IDO inhibitor, L­MT, as a potential novel therapeutic strategy for the treatment of melanoma.

Downregulation of RUNX3 moderates the frequency of Th17 and Th22 cells in patients with psoriasis.

Psoriasis is a common chronic inflammatory and T cell-meditated skin disease. Runt-related transcription factor 3 (RUNX3), one of the runt­domain family of transcription factors, has been reported to be a susceptibility gene for psoriasis. The present study was designed to delineate the role and underlying mechanism of RUNX3 involved in the differentiation of T helper (Th) 17 and Th22 cells in psoriasis. The results of the present study demonstrated that the expression of RUNX3 increased significantly in CD4­positive (CD4+) T cells from patients with psoriasis, compared with healthy controls. In addition, increased levels of interleukin (IL)­6, IL­20 and IL­22, and increased frequencies of Th17 and Th22 cells were found in the patients with psoriasis patients, compared with the healthy controls. It was also found that the overexpression of RUNX3 increased the levels of Th17­ and Th22­associated cytokines in the CD4+ T cells from the healthy controls. However, the inhibition of RUNX3 reduced the levels of the associated cytokines and decreased the frequency of Th17 and Th22 cells in the CD4+ T cells from the patients with psoriasis. Taken together, the present study suggested that RUNX3 regulated the differentiation of Th17 and Th22 cells in psoriasis, providing a promising therapeutic strategy for the treatment of psoriasis.

[Effects of Hedyotis diffusa Extract on Epidermal Growth Factor Induced Proliferation, Apoptosis, and TNF-α Induced Inflammatory Factors of HaCaT Cells].

Objective To observe the effects of Hedyotis diffusa extract (HDE) on the prolifera- tion, apoptosis, and inflammatory factors of HaCaT cells, and to explore its possible molecular mecha- nisms. Methods HaCaT cells were divided into 3 groups, the vehicle group, the control group, and 3 dose HDE groups. No epidermal growth factor (EGF) or HDE was added in the vehicle group. EGF was added with no HDE treatment in the control group. HDE (25, 50, 100 mg/mL) and EGF were added in the 3 HDE groups to co-culture HaCaT cells. The effects of HDE on EGF-induced proliferation of HaCaT cells were de- tected using CCK-8 assay. The effects of HDE on the growth cycle and apoptosis rate of HaCaT cells were measured using flow cytometry. Moreover, protein expression levels of Ki67, Bcl-xL, clAP1 , procaspase- 3, and cleaved caspase-3 were determined using Western blot. In addition, levels of IL-6, IL-8, and IL-10 in the supernatant were detected using ELISA. The level of phosphoration of NF-κB p65 (p-p65) was meas- ured using Western blot. Results Compared with the vehicle group, the absorbance of HaCaT cells and the expression level of Ki67 increased (P <0. 05, P <0. 01) ; levels of p-p65, IL-6, and IL-8 were elevated (P <0. 05, P <0. 01); IL-10 level was lowered (P <0.01) in the control group. Compared with the control group, the absorbance of HaCaT cells and the expression level of Ki67 decreased (P <0.05, P <0.01) ; levels of p-p65, IL-6, and IL-8 were reduced (P <0. 05, P <0. 01); IL-10 level was elevated (P <0. 05, P < 0.01) in the 3 dose HDE groups. Meanwhile, the apoptosis rate of HaCaT cells increased more in the 3 dose HDE groups than in the control group (P <0. 05, P <0. 01). The percentage of HaCaT cells at G1 phase was 58. 51 %, 73.12%, and 79. 95% in 25, 50, and 100 mg/mL HDE groups, respectively, showing statisti- cal difference when compared with that in the control group (52. 85%; P <0. 05, P <0. 01). The percentage and apoptosis rate of HaCaT cells at G1 phase were elevated more in 50 and 100 mg/mL HDE groups than in 25 mg/mL HDE group (P <0. 01). Besides, the percentage and apoptosis rate of HaCaT cells at G1 phase were elevated more in 100 mg/mL HDE group than in 50 mg/mL HDE group (P <0. 05). Compared with the control group, protein expression levels of Bcl-xL and cIAP1 were reduced in 100 mg/mL HDE group (P < 0. 01). There was no statistical difference in caspase-3 total amount (P >0. 05), but cleaved caspase-3 ex- pression increased with statistical difference (P <0. 01). Conclusion HDE inhibited the proliferation of HaCaT cells possibly by arresting HaCaT cell growth at G1 phase, promoted the apoptosis of HaCaT cells by stressing protein expressions of Bcl-xL and cIAPI , and elevating protein expressions of cleaved caspase-3, and suppressed inflammatory response of HaCaT cells via regulating NF-κB expression.

MicroRNA-138 regulates the balance of Th1/Th2 via targeting RUNX3 in psoriasis.

Psoriasis is a common chronic inflammatory and T cell-meditated autoimmune skin disease. A recent study found that Runt-related transcription factor 3 (RUNX3) is a susceptibility gene for psoriasis; however, its biological role in the disease has not been studied. RUNX3 was predicted to be the target gene of microRNA-138 (miR-138). The current research was designed to delineate the mechanism of miR-138 in regulating psoriasis via targeting RUNX3. In this study, we found that the expression of RUNX3 is increased significantly while the expression of miR-138 decreased significantly in CD4(+) T cells from psoriasis patients. Moreover, the luciferase report confirmed the targeting reaction between miR-138 and RUNX3. After transfection with the miR-138 inhibitor into CD4(+) T cells from healthy controls, we found that the inhibition of miR-138 increases RUNX3 expression and increased the ratio of Th1/Th2. Furthermore, the miR-138 mimic was transfected into CD4(+) T cells from psoriasis patients. The results showed that the overexpression of miR-138 inhibits RUNX3 expression and decreased the ratio of Th1/Th2 in CD4(+) T cells. Taken together, our study suggests that increased miR-138 regulates the balance of Th1/Th2 through inhibiting RUNX3 expression in psoriasis, providing a potential therapeutic target for psoriasis.

Effects of PTTG down-regulation on proliferation and metastasis of the SCL-1 cutaneous squamous cell carcinoma cell line.

AIMS: To study effects of down-regulation of pituitary tumor-transforming gene (PTTG) on proliferation and metastasis ability of the SCL-1 cutaneous squamous cell carcinoma (CSCC) cell line and explore related mechanisms. METHODS: SCL-1 cells were divided into 3 groups (untreated, siRNA control and PTTG siRNA). Cell proliferation assays were performed using a CCK-8 kit and proliferation and metastasis ability were analyzed using Boyden chambers. In addition, expression of MMP-2 and MMP-9 was detected by r-time qPCR and Western blotting. RESULTS: Down-regulation of PTTG could markedly inhibit cell proliferation in SCL-1 cells, compared to untreated and control siRNA groups (P < 0.05). Real-time qPCR demonstrated that expression levels of PTTG, MMP-2 and MMP-9 in the PTTG siRNA group were 0.8%, 23.2% and 21.3% of untreated levels. Western blotting revealed that expression of PTTG, MMP-2 and MMP-9 proteins in the PTTG siRNA group was obviously down-regulated. The numbers of migrating cells (51.38 ± 4.71) in the PTTG siRNA group was obviously lower than that in untreated group (131.33 ± 6.12) and the control siRNA group (127.72 ± 5.20) (P < 0.05), suggesting that decrease of proliferation and metastasis ability mediated by PTTG knock-down may be closely correlated with down-regulation of MMP-2 and MMP-9 expression. CONCLUSION: Inhibition of PTTG expression may be a new target for therapy of CSCC.

Scorpion venom component III inhibits cell proliferation by modulating NF-κB activation in human leukemia cells.

Scorpion venom contains various groups of compounds that exhibit anticancer activity against a variety of malignancies through a poorly understood mechanism. While the aberrant activation of nuclear factor κB (NF-κB) has been linked with hematopoietic malignancies, we hypothesized that scorpion venom mediates its effects by modulating the NF-κB signaling pathway. In the present study, we examined the effects of scorpion venom component III (SVCIII) on the human leukemia cell lines THP-1 and Jurkat and focused on the NF-κB signaling pathway. Our results showed that SVCIII inhibited cell proliferation, caused cell cycle arrest at G1 phase and inhibited the expression of cell cycle regulatory protein cyclin D1 in a dose-dependent manner in THP-1 and Jurkat cells. SVCIII also suppressed the constitutive NF-κB activation through inhibition of the phosphorylation and degradation of IκBα. NF-κB luciferase reporter activity was also inhibited by SVCIII. Our data suggest that SVCIII, a natural compound, may exert its antiproliferative effects by inhibiting the activation of NF-κB and, thus, has potential use in the treatment of hematopoietic malignancies, alone or in combination with other agents.

[Effect of blockade of NF-kappaB signaling pathway on cell apoptosis in cutaneous squamous cell carcinoma].

OBJECTIVE: To evaluate the siRNA-mediated inhibitory effect of nuclear factor-kappaB (NF-kappaB) p65 on expression of p65, and explore the effect of blockade of NF-kappaB signal pathway on cell apoptosis in cutaneous squamous cell carcinoma (cutaneous SCC). METHODS: Cutaneous SCC cell line SCL-1 cells were transfected with 50 nmol/L p65 siRNA. The expression level of p65 mRNA was measured using RT-PCR method at 0, 24, 48 and 72 h . Expressions of p65, bcl-2 and bax proteins were determined using Western blotting. Activities of caspase-3/9 was detected by Caspase-Glo®-3/7, 8 and 9 kit. Finally, cell apoptosis was detected using flow cytometry. RESULTS: The expression level of p65 mRNA in Cutaneous SCC SCL-1 cells was obviously down-regulated 48 h after transfection with p65 siRNA, and a significant difference was detected, as compared with 0 h after (0.23 ± 0.10 vs. 0.66 ± 0.05, P<0.05). The protein levels of p65 and bcl-2 decreased, and the bax protein level and activities of caspase-3/9 increased after transfection with p65 siRNA at h 48 . Further, the results of flow cytometry demonstrated that p65 siRNA could induce apoptosis of SCL-1 cells, and cell apoptosis ratio (20.28% ± 1.87%) in p65 siRNA group was significantly higher than that in the untreated group and control siRNA group (9.13% ± 1.51% and 9.37% ± 1.38%, respectively, F=47.532, P<0.01). CONCLUSION: p65 siRNA can block NF-kappaB signal pathway, down-regulate expression of bcl-2, elevate the bax level and increase the activities of caspase-3/9, suggesting that NF-kappaB signal pathway may be a key molecular target for therapy of cutaneous SCC.

Restoration of fragile histidine triad (FHIT) expression inhibits cell growth and induces apoptosis in cutaneous T-cell lymphoma cell line.

To investigate the effects of fragile histidine triad (FHIT) restoration on cell proliferation and apoptosis in human cutaneous T-cell lymphoma cell line, Hut-78, in vitro and in nude mouse. Wild-type FHIT gene was transfected by liposome into the Hut-78 cells. The alteration of cell growth curve and soft agar colony formation was studied in vitro and in nude mice. The FHIT gene was stably expressed in Hut-78 cells after the transfection. Compared with the controls, restoration of FHIT expression inhibited cell growth and induced apoptosis. Tumor formation in vivo was strongly suppressed by FHIT gene restoration. Our data demonstrate restoration of FHIT gene expression inhibit the tumor cell growth and provide an option for the treatment.