A Phenylurea Cytokinin, CPPU, Elevated Reducing Sugar and Correlated to Andrographolide Contents in Leaves of Andrographis paniculata (Burm. F.) Wall. Ex Nees.

Cytokinins are phytohormones that play multiple roles to control plant growth and development. In this study, leaf biomass and the production of andrographolide compounds in a medicinal plant Andrographis paniculata were significantly increased after exogenously treating with the synthetic cytokinin cytokinin-1-(2-chloro-4-pyridyl)-3-phenylurea (CPPU) at 0 (water), 5, or 10 mg L-1 and observed the results for 24 h, 48 h, and 7 days of treatment. It was found that CPPU could significantly enhance new axillary bud formation and further promote branching 4.6-5.6-fold higher, resulting in higher fresh weight (FW) and dry weight (DW) than the control. Application of CPPU at 5 mg L-1 significantly promoted the highest contents of total reducing sugar at 2.5-fold in leaves and at 1.5-fold in roots. Although treatments of CPPU significantly affected the increasing contents of chlorophyll and carotenoid (1.2-1.6-fold), CPPU at 10 mg L-1 slightly caused leaf stress and chlorophyll reduction. Interestingly, 5 mg L-1 CPPU could enhance andrographolide content, an active anti-infectious compound in Andrographis paniculata (2.2-fold higher than the control) that reached the highest content at 24 h after treatment. This study suggested that CPPU should be suitable for field application to promote leaf yields and induce the production of useful pharmaceutical compounds in Andrographis paniculata.

Auxin-cytokinin interaction and variations in their metabolic products in the regulation of organogenesis in two Eucomis species.

In the current study, we evaluated the effect of α-naphthaleneacetic acid (NAA) individually or in combination with different cytokinins (CKs) including benzyladenine (BA), meta-topolin (mT) and isopentenyladenine (iP) on organogenesis, auxin and CK content in Eucomis autumnalis subspecies autumnalis (EA) and Eucomis zambesiaca (EZ). These species were used as model plants due to their ornamental and medicinal properties. Three leaf explants were inoculated in screw-cap jars containing 30mL Murashige and Skoog (MS) media supplemented with 5µM NAA alone or in combination with 5µM CK (BA, mT or iP). After 10 weeks (EA) or 15 weeks (EZ), parameters including shoot and root growth as well as plant fresh weight were recorded. For analysis of auxin and CK content, whole plantlets were harvested, pooled and freeze-dried for the different treatments. In both species, shoot and root proliferation as well as plant biomass were generally higher when NAA was combined with the individual CK than in NAA or CK treatment. The highest concentration of indole-3-acetic acid (IAA, 619pmolg-1 DW) and 2-oxindole-3-acetic acid (OxIAA, 2381pmolg-1 DW) were observed in EA-treated with NAA alone while mT treatment (without NAA) had the most abundant indole-3-acetyl-l-aspartic acid (IAAsp, 904 and 582pmolg-1 DW for EA and EZ, respectively) in both species. A significant concentration of total endogenous CK accumulated in both Eucomis regenerants from mT and mT+NAA when compared to the other treatments. The majority of the detected CKs were of the aromatic CK-type, mainly free bases. The potential physiological roles of these quantified phytohormones in relation to the observed morphological responses are discussed.

Unraveling the intricate nexus of molecular mechanisms governing rice root development: OsMPK3/6 and auxin-cytokinin interplay.

The root system is an imperative component of a plant, involved in water and nutrient acquisition from the soil. Any subtle change in the root system may lead to drastic changes in plant productivity. Both auxin and cytokinin are implicated in regulating various root developmental aspects. One of the major signaling cascades facilitating various hormonal and developmental allocations is the Mitogen Activated Protein Kinase (MAPK) cascade. Innumerable efforts have been made to unravel the complex nexus involved in rice root development. In spite of a plethora of studies, a comprehensive study aiming to decipher the plausible cross-talk of MAPK signaling module with auxin and cytokinin signaling components in rice is missing. In the present study, extensive phenomics analysis of different stages of rice roots; transcript profiling by qRT-PCR of entire gene family of MAPK, MAPKK and PIN genes; as well as protein level and activity of potential MAPKs was investigated using western and immuno kinase assays both on auxin and cytokinin treatment. The above study led to the identification of various novel rice root specific phenotypic traits by using GiA roots software framework. High expression profile of OsMPK3/6, OsMKK4/5 and OsPIN 1b/9 and their marked transcript level modulation in response to both auxin and cytokinin was observed. Finally, the protein levels and activity assay further substantiated our present findings. Thus, OsMPK3/6-OsMKK4/5 module is elucidated as the putative, key player in auxin-cytokinin interaction augmenting their role by differentially regulating the expression patterns of OsPIN 1b/9 in root development in rice.

Combination of cytokinin and auxin induces apoptosis, cell cycle progression arrest and blockage of the Akt pathway in HeLa cells.

Plant cytokinins and auxins have recently been proposed as novel cancer therapies, which proceed via different mechanisms; however, their combined use has not been investigated. To the best of our knowledge, the present study was the first to show that the cytokinin ortho-methoxytopolin-riboside (MeoTR) strongly inhibited the proliferation of HeLa cells, the effect of which was synergistically enhanced by auxin indole-3-acetic acid (IAA), while IAA demonstrated to have no cytotoxic effects on cells. MeoTR was found to activate intrinsic and extrinsic caspase-dependent pathways, and IAA potentiated this activation. In addition, these effects were blocked by Z-Val-Ala-Asp-fluoromethylketone (Z-VAD-FMK), a pan-specific-caspase-inhibitor. IAA increased the MeoTR- induced inhibition of B cell lymphoma 2 (Bcl-2) and survivin, whereas IAA-only decreased Bcl-2 expression. MeoTR downregulated phosphorylated (p)-pyruvate dehydrogenase kinase 1, p-Akt and p-glycogen synthase kinase 3ß, the effect of which was more potent in combination with IAA, despite the weak effect of IAA alone. LY294002, an Akt-inhibitor, was able to increase the inhibition of p-Akt through MeoTR and combination treatment. IAA and MeoTR increased the proportion of cells in S phase independently. However, the combination treatment induced a further increase. In addition, IAA and MeoTR treatment downregulated protein levels of cyclin A, cyclin-dependent kinase 2 (CDK2) and p-CDK2, and upregulated protein levels of p21 and p27. Furthermore, the combination treatment enhanced these effects, indicating that IAA potentiated the inhibitory effect of MeoTR on HeLa cells via cell cycle progression arrest and accumulation in S phase, coupled with the negative regulation of Bcl-2. In conclusion, the results of the present study suggested that treatment with these two phytohormones in combination, may offer a novel therapeutic strategy for the treatment of malignant cervical cancer.

Concentrações de BAP sobre a proliferação in vitro de brotos de Lippia alba [(Mill. )N. E. Brown] / BAP concentrations on in vitro proliferation of Lippia alba [(Mill. ) N. E. Brown] shoots

Lippia alba é uma planta de origem brasileira, pertencente à família Verbenaceae. É conhecida por combater a insônia e a asma, além de possuir ação anticonvulsivante e antifúngica. A micropropagação tem sido utilizada para multiplicação de várias espécies com propriedades medicinais, e o BAP é a citocinina sintética mais utilizada para a multiplicação in vitro. O objetivo deste trabalho foi estabelecer a concentração adequada de BAP na multiplicação in vitro de L. alba. Explantes constituídos de segmentos nodais provenientes de plântulas já estabelecidas in vitro com aproximadamente 0,5 cm foram inoculados em meio MS, suplementado com 0,0; 0,5; 1,0 e 1,5 mg L-1 de BAP em adição de 30 g L-1 de sacarose. A utilização de 1,5 mg L-1 de BAP promove a multiplicação in vitro de L. alba. A ausência deste regulador propicia maior número de folhas e o uso na concentração de 0,5 mg L-1 aumentou as massas fresca e seca de parte aérea nesta espécie.

Lippia alba is a Brazilian plant, belonging to the Verbenaceae family. It is known to combat insomnia and asthma, also has antifungal and anticonvulsant actions. Micropropagation has been used for propagation of various species with medicinal properties, and BAP is the most widely used synthetic cytokinin for in vitro multiplication. The aim of this study was to determine the appropriate concentration of BAP on in vitro multiplication of L. alba. Explants consisting of nodal segments from seedlings already established in vitro with approximately 0.5 cm were inoculated on MS medium supplemented with 0; 0.5; 1.0 and 1.5 mg L-1 of BAP and 30 g L-1 of sucrose. The use of 1.5 mg L-1 of BAP promotes the in vitro multiplication of L. alba. The absence of this regulator provides a greater number of leaves and its use at the concentration of 0.5 mg L-1 increased shoots' fresh and dry weights in this specie.

Cálcio e fitorreguladores no desenvolvimento e estado nutricional de Mentha spicata x suaveolens cultivada em solução nutritiva / Calcium and plant growth regulators on the development and nutritional status of Mentha spicata x suaveolens cultivated in nutrient solution

O trabalho teve como objetivo estudar o desenvolvimento e o estado nutricional de Mentha spicata x suaveolens cultivada em diferentes níveis de Calcio em solução nutritiva com aplicação de fitorreguladores. O experimento constou de esquema fatorial (4x4) sendo quatro níveis de Ca2+ na solução nutritiva (160, 120, 80 e 40 mg L-1) e três aplicações foliares, com intervalos de 15 dias, de ácido naftaleno acético (NAA), ácido giberélico (GA3), benzinaladenina (BA) e água destilada como testemunha. As plantas foram coletadas 60 dias após a transferência para a solução e avaliadas as variáveis, matéria fresca e seca, área foliar e número de folhas; volume, comprimento e superfície radiculares e teores de Ca, K, Mg, N e P das folhas. As variáveis foram submetidas à análise de variância e as médias comparadas pelo teste Tukey. Não foram encontradas diferenças entre os níveis de Ca para a produção de biomassa. A auxina não influenciou a matéria seca das plantas, embora tenha aumentado os teores foliares de Ca; a citocinina acarretou diminuição da matéria seca total e a giberelina aumento da matéria seca do caule. A variação de Ca2+ não influenciou os teores foliares de N, P, K e Mg. Houve resposta para aplicação de Ca2+, com teores foliares atingindo o máximo de 15,73 mg kg-1 na dose de 137,81 mg L-1 pela análise de regressão

This study aimed to verify the development and the nutritional status of Mentha spicata x suaveolens cultivated under different Ca2+ levels in nutrient solution with application of plant growth regulators. The experiment was in factorial design (4X4) with four Ca2+ levels in the nutrient solution (160, 120, 80 and 40 mg L-1) and three leaf applications, at 15-day intervals, of naphthalene acetic acid (NAA), gibberellic acid (GA3), benzyladenine (BA) and distilled water as control. Plants were harvested at 60 days after transferring to the nutrient solution and the following variables were evaluated fresh and dried matter, leaf area, number of leaves, volume, length and surface of roots, and leaf content of Ca, K, Mg, N and P. The variables were subjected to analysis of variance and means compared by Tukey's test. There were no differences between Ca levels for biomass production. The auxin did not influence the dry matter of plants although it increased the leaf content of Ca2+, the cytokinin reduced total dry matter and the gibberellin increased the stem dry matter. The change in Ca2+ did not influence the leaf content of N, P, K and Mg. There was response for the application of Ca2+, reaching the maximum of 15.73 mg L-1 leaf content at the level of 137.81 mg L-1, according to regression analysis

Citocininas na multiplicação in vitro de hortelã-pimenta (Mentha x Piperita L. ) / Cytokinins on in vitro multiplication of peppermint (Mentha x Piperita L. )

O hortelã-pimenta é um hibrido triplo, Mentha x piperita, utilizado de forma medicinal no tratamento de náuseas, cólicas gastrointestinais, flatulências, cálculos biliares, icterícia, ansiedade, expectoração e expulsão de vermes intestinais. Na micropropagação, o BAP (6-benzilaminopurina) e KIN (cinetina) têm sido as fontes de citocininas mais empregadas. O objetivo deste trabalho foi avaliar o efeito de reguladores vegetais na multiplicação in vitro de hortelã-pimenta. Explantes constituídos de segmentos nodais provenientes de plântulas já estabelecidas in vitro com aproximadamente 0,5 cm foram inoculados em meio MS, suplementado com diferentes associações de BAP e KIN, em adição de 30 g L-1 de sacarose. As concentrações de citocininas utilizadas foram 0,0; 1,0 e 2,0 mg L-1 de BAP e 0,0; 0,5 e 1,0 mg L-1 de KIN. O delineamento experimental foi o inteiramente casualizado, com nove tratamentos consistindo de cinco frascos cada um, sendo que cada frasco continha quatro explantes. A utilização de 2,0 mg L-1 de BAP promove a multiplicação in vitro de hortelã-pimenta, porém, diminui o índice de sobrevivência. O uso de citocininas aumenta a massa fresca e seca dos explantes e a ausência destes reguladores propicia o alongamento nesta espécie.

Peppermint is used for medical treatments of nausea, gastrointestinal cramps, flatulence, gallstones, jaundice, anxiety, sputum and expulsion of intestinal worms. In micropropagation, BAP (6-benzylaminopurine) and KIN (kinetin) have been the most widely used sources of cytokinins. The aim of this study was to evaluate the effect of plant growth regulators in multiplication of peppermint. Explants consisting of nodal segments from seedlings already established in vitro with approximately 0,5 cm were inoculated on MS medium supplemented with different combinations of BAP and KIN and 30 g L-1 of sucrose. The concentrations of cytokinins used were 0,0; 1,0 and 2,0 mg L-1 of BAP and 0,0; 0,5 and 1,0 mg L-1 of KIN. The experiment was set up in a completely randomized design with nine treatments consisting of five bottles each, and each vial contained four explants. The use of 2,0 mg L-1 of BAP promotes the in vitro multiplication of peppermint, but decreases the survival rate. Cytokinins increase fresh and dry weight of explants and the absence of these regulators provides the elongation in this specie.

Long-term efficacy and safety of topical PRK 124 (0.125%) lotion (Pyratine-XR) in the treatment of mild-to-moderate rosacea.

BACKGROUND: Many patients with rosacea cannot tolerate extended treatment periods with topical agents because their skin sensitivity is often increased. OBJECTIVE: To determine the long-term efficacy and tolerability of a new moisturizing lotion for improving the signs and symptoms of mild-to-moderate rosacea. METHODS: In a 48-week, open-label study, a moisturizing lotion containing furfuryl tetrahydropyranyladenine as PRK-124 (0.125%, Pyratine-XR, Senetek PLC, Napa, CA) was applied twice daily by 18 subjects with mild-to-moderate rosacea. Clinical improvements were assessed by the treating physician. Skin barrier function was measured by transepidermal water loss after treatment. Tolerability and cosmetic outcome were evaluated by subjects. RESULTS: Subjects experienced a mean 44 percent reduction in erythema severity and a mean 89 percent reduction in inflammatory lesion count at week 48. Reductions were significant (P < 0.05) in both erythema and lesions at weeks 24, 36 and 48. Statistically significant (P < or = 0.05) improvements in telangiectasias, transepidermal water loss and dryness were noted. Overall clinical improvement was observed in 81 percent of subjects and the investigator's global assessment steadily improved throughout the study. Treatments were well-tolerated and cosmetically acceptable. Treatment-induced skin irritation was not observed. CONCLUSION: The new moisturizing lotion containing furfuryl tetrahydropyranyladenine as PRK 124 is efficacious, does not irritate skin, and is well tolerated for at least 48 weeks.

Topical PRK 124 (0.125%) lotion for improving the signs and symptoms of rosacea.

BACKGROUND: Current treatments for acne rosacea are often associated with unsatisfactory outcomes and adverse effects. OBJECTIVE: To determine the efficacy and tolerability of a new moisturizing lotion for improving the clinical signs and symptoms of mild-to-moderate acne rosacea. METHODS: In a 12-week, open-label study, a moisturizing lotion containing furfuryl tetrahydropyranyladenine as PRK124 (0.125%, Pyratine-XR, Senetek PLC, Napa, CA) was applied twice daily to subjects with mild-to-moderate rosacea. Improvement in the appearances of erythema and papules were assessed by the treating physician. Skin barrier function was measured by transepidmal water loss after treatment. Tolerability and cosmetic outcome were evaluated by patients. RESULTS: Twenty-one participants completed the study. Overall clinical improvement was observed in 80% of subjects, with most showing mild-to-moderate improvement. Erythema, papule counts, and telangiectasia were reduced. The reduction in TEWL was significant at weeks 4 (p = 0.01), 8 (p < 005), and 12 (p < 0.001). Rosacea symptoms (burning, stinging, dryness) were progressively reduced, with reduction in dryness achieving statistical significance at weeks 4 (p = 0.035), 8 (p = 0.037) and 12 (p = 0.016). Treatments were well tolerated and cosmetic outcomes were acceptable. Treatment-induced irritation was not observed. CONCLUSION: The new moisturizing lotion containing furfuryl tetrahydropyranyladenine as PRK124 shows a continued trend toward improvement of skin barrier function and the appearances of erythema and papules associated with mild-to-moderate rosacea during 12 weeks of treatment.

Effect of a metabotropic glutamate receptor 5 antagonist, MPEP, on the nociceptive response induced by intrathecal injection of excitatory aminoacids, substance P, bradykinin or cytokines in mice.

Metabotropic glutamate receptors (mGluRs) are expressed abundantly in the spinal cord and have been shown to play important roles in the modulation of nociceptive transmission and plasticity. In this study, the involvement of metabotropic glutamate receptor 5 (mGluRs) in the nociceptive response induced by intrathecal injection (i.t.) of excitatory aminoacids, substance P (SP), bradykinin (BK) and cytokines in mice was demonstrated. The administration of 2-methyl-6-(phenylethynyl)-pyridine (MPEP; 10-50 nmol/site, i.t.) caused a significant inhibition in the nociceptive response induced by glutamate and trans-ACPD with maximal inhibitory effects of 36 +/- 7% and 56 +/- 5%, respectively. MPEP completely failed to affect the nociception induced by alpha-amino-3-hydroxy-5-mehtyl-4-isoxazolepropionic acid (AMPA; 135 pmol/site), kainate (110 pmol/site) and N-methyl-D-aspartate (NMDA; 450 pmol/site). MPEP also reduced the nociceptive response induced by SP (135 ng/site, i.t.), BK (0.1 microg/site), tumor necrosis factor-alpha (TNF-alpha; 0.1 pg/site) and interleukin-1beta (IL-1beta; 1 pg/site) with maximal inhibitions of 29 +/- 5%, 37 +/- 5%, 83 +/- 3% and 88 +/- 1%, respectively. Together, these results indicate the involvement of mGluRs, more specifically of subtype-5, in the nociceptive response induced by i.t. injection of excitatory aminoacids, SP, BK and cytokines in mice.

A clinical study of topical Pyratine 6 for improving the appearance of photodamaged skin.

OBJECTIVE: Pyratine 6 has been shown to have antiaging effects in human skin cells. The purpose of this study is to determine the cosmetic efficacy and tolerance of topical Pyratine 6 (0.10%) over 12 weeks for improving the baseline clinical signs and symptoms of photodamaged facial skin. METHODS: A single-arm longitudinal study with observations at 2, 4, 8, and 12 weeks was conducted. Forty healthy women with mild to moderate signs of photodamaged facial skin applied Pyratine 6 twice daily for 12 weeks. Efficacy and safety were evaluated by clinical observations, digital photography, transepidermal water loss (TEWL), skin capacitance, and silicon replicas at each time point. RESULTS: Topical Pyratine 6 achieved significant improvement from baseline in roughness and skin moisture content after 2 weeks. After 4 weeks, significant improvement in fine wrinkles, mottled hyperpigmentation, and TEWL were observed. Improvements in most parameters were maintained throughout the remaining weeks of the study. For most silicon replica parameters, changes were consistent with increased skin smoothing. Facial erythema was reduced at 2 weeks and further reduced at 4, 8, and 12 weeks. Adverse effects were minimal and transient. CONCLUSIONS: Treatment with Pyratine 6 over 12 weeks improves roughness and skin moisturization in 2 weeks compared to baseline and mottled hyperpigmentation and fine wrinkles in 4 weeks compared to baseline. Reduction in facial erythema occurs as early as 2 weeks. Adverse effects are minimal and transient.