Targeting CXCL5 in Pancreatic Cancer Cells Inhibits Cancer Xenograft Growth by Reducing Proliferation and Inhibiting EMT Progression.

BACKGROUND: Pancreatic cancer (PC) is the most lethal malignant tumor, with average survival period of about 10 months. C-X-C ligand 5 (CXCL5), an important chemokine for immune cell accumulation in tumor tissues, has been reported to be involved in a variety of human cancers. However, the exact role of CXCL5 in PC progression has not been well defined. METHODS: The expression of CXCL5 in PC was analyzed based on online databases and clinical specimens immunohistochemical staining, and Western blotting of CXCL5 in PC cell lines and patient samples. The correlation between CXCL5 expression and prognosis in PC was explored. The role of CXCL5 in PC was investigated through in vitro and in vivo experiments. RESULTS: The expression of CXCL5 was significantly increased in PC tissues compared with that in pancreas tissues, and CXCL5 high expression predicts poor prognosis in PC patients. Further analyses demonstrated that overexpression of CXCL5 in PC cells was positively related to higher proliferation rate, higher migration ability, and higher EMT markers including SNAI2 and TWIST1 of tumor cells in vitro. Consistently, the knockdown of CXCL5 in PC cells harmed the proliferation rate, migration ability, and expression of EMT indexes of tumor cells in vitro. Importantly, knockdown of CXCL5 inhibited the growth of xenograft tumors in vivo. CONCLUSION: CXCL5 high expression predicts poor prognosis in PC patients. CXCL5 promotes PC cell growth and EMT process. Inhibition of CXCL5 may be a potential therapeutic approach for PC.

Paraneoplastic neurological syndrome caused by cystitis glandularis: A case report and literature review.

BACKGROUND: Paraneoplastic neurological syndrome (PNS) is an unusual event. PNS caused by cystitis glandularis (CG) or a bladder tumor is extremely rare; hence, missed diagnosis or misdiagnosis can easily occur. To date, approximately 21 cases have been reported in PubMed. CASE SUMMARY: We report a case of PNS caused by CG and describe the clinical and imaging features. The main clinical feature was advanced cognitive impairment, and early clinical features were memory impairment, decreased computational ability, and abnormal behavior. Later clinical features were dementia, vomiting, inability to eat and walk, urinary incontinence, and hematuria. Imaging features on cranial magnetic resonance imaging were diffuse white matter lesions. Paraneoplastic tumor markers were normal. A total abdominal computed tomography scan showed multiple thickened areas on the bladder wall with local prominence. Cystoscopy revealed a volcanic protuberance on the posterior wall of the bladder with a diameter of 6 cm and no pedicle. The postoperative pathological diagnosis was CG. The patient recovered well following resection of CG. PNS cases caused by previous bladder tumors can be retrieved from PubMed to describe the clinical signs and prognosis of PNS. CONCLUSION: The main clinical feature of PNS caused by CG was dementia, and the imaging features were diffuse cerebral white matter lesions. Resection of CG lesions is the fundamental treatment for PNS induced by CG. This case highlights the importance of etiological treatment.

Architectural effects regulate resource allocation within the inflorescences with nonlinear blooming patterns.

PREMISE: Spatial and temporal resource allocations within inflorescences have been well-studied in many plants based on flowering sequence or floral position. However, there had been few attempts to investigate architectural effects and resource competition in species where the blooming pattern does not follow a linear positional pattern within the inflorescence. Moreover, most flowering plants show female-biased sex allocation in early or basal flowers, but it is unclear in species with inherent and changeless ovule production. METHODS: We investigated intra-inflorescence variation in reproductive traits of Salvia przewalskii, a perennial herb with 4-ouvle ovary flowers and flowering sequence-floral position decoupled inflorescences. To detect the effects of resource competition and architectural effects on reproductive success, we manipulated inflorescence (removed floral buds by position and flowering sequence) and pollination (opened and supplemented pollination). RESULTS: Pollen production and dry mass deceased from bottom to top flowers but did not significantly differ following flowering sequence, resulting in male-biased sex allocation in basal flowers. The seed production, fruit set, and bud development exhibited significant declining trends from proximal to distal positions regardless of the thinning and pollen treatments. Meanwhile, the seed production, fruit set, and bud development success did not significant differ when thinning was conducted according to flowering sequence. CONCLUSIONS: Architectural effects plays a crucial role in resource allocation within decoupled flowering inflorescences. Moreover, our results highlighted that inherent floral traits such as changeless ovule production, may modify architectural effects on sex allocation.

Speciation along the elevation gradient: Divergence of Roscoea species within the south slope of the Himalayas.

The Himalayas with dramatic elevation gradient is one of the global biodiversity hotspots. Although origin of biodiversity of the Himalayas is of great concern, the speciation process within the Himalayas is poorly known. Roscoea within the Himalayas serve as a good model system to test the speciation process along an elevation gradient. 32,375 unlinked SNPs were used to reconstruct phylogenetic relationships and introgression analyses in D-statistics and Fastsimicoal2. Species distribution modeling (SDM) was used to simulate habitat shift of Roscoea species during climate changes. Phylogeny suggested that the speciation order, except R. capitata, was from highland to lowland. D-statistics analyses suggested significant bidirectional ancient introgression between elevation-neighboring clades but no introgression between R. capitata and othern clades and no introgression among extant species. Fastsimicoal2 suggested interspecific introgressions were asymmetric. SDM predicted that habitats of Roscoea shifted to low elevation during cooling age. These results suggested that the sudden uplift of the Himalayas likely promoted speciation by vicariance, and climate cooling drove species divergence towards lower elevation. This study provides explanations for the origin of biodiversity within the Himalayas, and an insight to understand speciation along elevation in the mountainous regions.

Chloroplast Genome Evolution in Four Montane Zingiberaceae Taxa in China.

Chloroplasts are critical to plant survival and adaptive evolution. The comparison of chloroplast genomes could provide insight into the adaptive evolution of closely related species. To identify potential adaptive evolution in the chloroplast genomes of four montane Zingiberaceae taxa (Cautleya, Roscoea, Rhynchanthus, and Pommereschea) that inhabit distinct habitats in the mountains of Yunnan, China, the nucleotide sequences of 13 complete chloroplast genomes, including five newly sequenced species, were characterized and compared. The five newly sequenced chloroplast genomes (162,878-163,831 bp) possessed typical quadripartite structures, which included a large single copy (LSC) region, a small single copy (SSC) region, and a pair of inverted repeat regions (IRa and IRb), and even though the structure was highly conserved among the 13 taxa, one of the rps19 genes was absent in Cautleya, possibly due to expansion of the LSC region. Positive selection of rpoA and ycf2 suggests that these montane species have experienced adaptive evolution to habitats with different sunlight intensities and that adaptation related to the chloroplast genome has played an important role in the evolution of Zingiberaceae taxa.

Sexual conflict in protandrous flowers and the evolution of gynodioecy.

Sexual interference between male and female function in hermaphrodite plants is reduced by protandry. In environments with insufficient pollinator service, prolongation of male function owing to limited pollen removal could restrict the duration of female function and lower seed production. We provide evidence that this form of sexual conflict has played a role in the spread of females in gynodioecious populations of Cyananthus delavayi in the pollen-limited environments in which this subalpine species occurs. Using field experiments involving artificial pollen removal from the strongly protandrous flowers of hermaphrodites, we demonstrated a trade-off between male- and female-phase duration with no influence on overall floral longevity. Pollen removal at the beginning of anthesis resulted in hermaphrodite seed production matching that of females. In contrast, restricted pollen removal increased the duration of male function at the expense of female function lowering maternal fertility compared to females. This pattern was evident in five populations with females experiencing a twofold average seed fertility advantage compared to hermaphrodites. Gynodioecy often appears to evolve from protandrous ancestors and pollen limitation is widespread in flowering plants suggesting that sexual conflict may play an unappreciated role in the evolution of this form of sexual dimorphism.

Functional implications of the specialized staminal appendages in alpine ginger (Roscoea spp.: Zingiberaceae).

Floral organs are widely believed to enhance the pollination and reproductive success of angiosperms. However, the functional implication of some floral structures is still unknown. In this study, we explored the functional role of staminal appendages on male and female reproductive success of Himalayan Roscoea spp. and tested if their function differed between species with biotic pollination and autonomous selfing. Phenotypic manipulation is a powerful approach to test the functional effect of a particular trait on plant fitness. We compared various proxies of pollination success between intact flowers and flowers with manually excised staminal appendages. We found that the rate of visitation did not differ between intact and manipulated flowers. Our results revealed that in outcrossing Roscoea spp., the staminal appendages act as triggering devices to facilitate pollen release and deposition and also to manipulate the foraging position of pollinators to ensure both male and female reproductive success. In contrast, in autonomously selfing Roscoea spp., the removal of staminal appendages did not affect any aspect of pollination processes. Our results suggest that the staminal appendages are an integral component of outcrossing in Roscoea spp. and are maintained by selection pressure through both male and female reproductive success. This study provides important insights on how variation in breeding systems can provoke changes in the structure and function of floral organs among congeners.

Elevation-related variation in the population characteristics of distylous Primula nivalis affects female fitness and inbreeding depression.

The population characteristics of distylous species are highly sensitive to stochastic natural selection pressure. Therefore, populations growing under different environmental conditions may vary in floral morph ratios, potentially affecting female fitness and leading to inbreeding depression. However, the variation in offspring quality among populations as a result of inbreeding depression is poorly understood in distylous species. This study investigates variations in plant density, seed mass, seed viability, female fitness, and post-dispersal inbreeding depression in both sexual morphs (long-styled and short-styled plants) of the distylous Primula nivalis that were subjected to different pollination treatments along an elevational gradient from 1657 to 2704 m a.s.l. Population characteristics (morph plant density and ratio) and fruit set were significantly affected by sexual morph and elevation. Plant density and fruit set frequencies were lower for short-styled than for long-styled plants at 2704 m a.s.l. The seeds from the cross-pollinated flowers of both morphs were higher in quality than those of self-pollinated flowers. The female fitness of seeds from cross-pollinated flowers of both morphs was higher than that of seeds from open-pollinated and self-pollinated flowers. The female fitness of seeds from long-styled flowers was higher than that of seeds from short-styled flowers at all elevations. Inbreeding depression increased with elevation among plants with short-styled flowers but not among those with long-styled flowers. Variation in the elevation-dependent mating system might influence female fitness and affect inbreeding depression in both floral morphs. In conclusion, the low quality of seeds from short-styled flowers at high elevations might decrease short-styled flower frequency, affecting population characteristics.

Water mediates fertilization in a terrestrial flowering plant.

Water-mediated fertilization is ubiquitous in early land plants. This ancestral mode of fertilization has, however, generally been considered to have been lost during the evolutionary history of terrestrial flowering plants. We investigated reproductive mechanisms in the subtropical ginger Cautleya gracilis (Zingiberaceae), which has two pollen conditions - granular and filiform masses - depending on external conditions. We tested whether rain transformed granular pollen into filiform masses and whether this then promoted pollen-tube growth and fertilization of ovules. Using experimental manipulations in the field we investigated the contribution of water-mediated fertilization to seed production. Rain caused granular pollen to form filiform masses of germinating pollen tubes, which transported sperm to ovules, resulting in fertilization and seed set. Flowers exposed to rain produced significantly more seeds than those protected from the rain, which retained granular pollen. Insect pollination made only a limited contribution to seed set because rainy conditions limited pollinator service. Our results reveal a previously undescribed fertilization mechanism in flowering plants involving water-mediated fertilization stimulated by rain. Water-mediated fertilization is likely to be adaptive in the subtropical monsoon environments in which C. gracilis occurs by ensuring reproductive assurance when persistent rain prevents insect-mediated pollination.

Pollinator-mediated selection on floral traits varies in space and between morphs in Primula secundiflora.

Elucidating how variation in selection shapes the evolution of flowers is key to understanding adaptive differentiation processes. We estimated pollinator-mediated selection through female function in L-morph (long-style and short-anther phenotype) and S-morph (short-style and long-anther phenotype) flowers among four Primula secundiflora populations with different pollinator assemblages. Variation in pollinator assemblage strongly contributed to differences in reproductive success among populations and between morphs of the primrose species. A wider corolla tube width was selected in the bumblebee-dominated populations, whereas shorter corolla tube length and wider corolla tube width were selected in the syrphid fly-dominated populations. Morph-specific variation in pollinator-mediated selection on corolla tube length was detected in the syrphid fly-dominated populations. A shorter corolla tube was selected in the L-morph flowers. However, similar selective pressure on this trait was not observed in the S-morph flowers. These results show that variation in pollinator assemblage leads to variation in selection in space and between morphs. The findings highlight the potential forces of different pollinator agents in driving floral evolution in this primrose species.

Reproductive isolation in alpine gingers: How do coexisting Roscoea (R. purpurea and R. tumjensis) conserve species integrity?

Multiple barriers may contribute to reproductive isolation between closely related species. Understanding the relative strength of these barriers can illuminate the ecological factors that currently maintain species integrity and how these factors originally promoted speciation. Two Himalayan alpine gingers, Roscoea purpurea and R. tumjensis, occur sympatrically in central Nepal and have such similar morphology that it is not clear whether or how they maintain a distinct identity. Our quantitative measurements of the components of reproductive isolation show that they are, in fact, completely isolated by a combination of phenological displacement of flowering, earlier for R. tumjensis and later for R. purpurea, and complete fidelity of visitation by different pollinator species, bumblebees for R. tumjensis and a long-tongued fly for R. purpurea. Furthermore, the nectar of R. tumjensis flowers is available to the shorter tongued bumblebees while R. purpurea nectar is less accessible, requiring deep probing from long-tongued flies. Although flowering phenology is a strong current barrier that seemingly obviates any need for pollinator discrimination, this current pattern need not reflect selective forces occurring at the initial divergence of R. tumjensis. There has been considerable pollinator switching during the radiation of the Himalayan Roscoea, and the association of flowering time with type of pollinator in these sympatric species may have originated among the earliest or latest flowering individuals or populations of an ancestor to exploit either bumblebee activity early in the breeding season or long-tongued fly abundance later in the season. These two sympatric Roscoea species add to accumulating evidence of the primacy of prezygotic pollination traits in speciation among angiosperms even in the absence of postzygotic incompatibility.

Variation in the degree of reciprocal herkogamy affects the degree of legitimate pollination in a distylous species.

Distyly is a widespread floral polymorphism characterized by the flowers within a population showing reciprocal placement of the anthers and stigma. Darwin hypothesizes that distyly evolves to promote precise pollen transfer between morphs. Primula chungensis exhibits two types of anther heights, and these two types of anthers show pollen of two different size classes. To understand whether the stigma could capture more pollen grains from the anthers of the pollen donor as the separation between the stigma of pollen receiver and the anther of pollen donor decreased, the present research assessed the source of the pollen load in a series of open-pollinated flowers with continuous variation of style lengths. Individuals with continuous variation of style length were tagged, and the selected flowers in the tagged plants were emasculated the day before dehiscence. The stigma of the emasculated flowers was fixed in fuchsin gel at the end of blooming. We assessed the pollen sources on each stigma by taking photos under a microscope and measured the diameter of each conspecific pollen grain with ImageJ. We found that a shorter distance from the stigmas to the anthers of a pollen donor gave the flower a higher capacity to receive pollen from those anthers. Our result provides a new evidence that distyly could promote the pollen transfer between morphs, which is consistent with Darwin's hypothesis of disassortative pollination. An alternative hypothesis for the evolution of distyly (e.g. selfing avoidance) might also be true, but less likely, because self-incompatibility would greatly avoid self-fertilization for many distylous species.

Correlation between the timing of autonomous selfing and floral traits: a comparative study from three selfing Gentianopsis species (Gentianaceae).

About 20% of angiosperms employ self-fertilization as their main mating strategy. In this study, we aimed to examine how the selfing timing correlated with floral traits in three Gentianopsis species in which autonomous selfing is achieved through filament elongation. Although the three Gentianopsis species exhibit no significant variation in their capacity for autonomous selfing, flowers of G. grandis last longer, are larger and have a higher corolla biomass, P/O ratios and male biomass allocation than those of G. paludosa, and especially those of G. contorta. Autonomous selfing occurs in the early floral life of G. paludosa and G. contorta and in the later floral life of G. grandis. Seed production mainly results from autonomous selfing in G. paludosa and G. contorta; however, G. grandis could be more described as having a mixed mating system. We suggest that autonomous selfing in later floral life increases the chance of cross-pollination prior to this, while autonomous selfing in early floral life offers a selective advantage to plants by reducing the resource investment in traits that may increase pollinator attraction and visitation.

Phenotypic selection on flowering phenology and pollination efficiency traits between Primula populations with different pollinator assemblages.

Floral traits have largely been attributed to phenotypic selection in plant-pollinator interactions. However, the strength of this link has rarely been ascertained with real pollinators. We conducted pollinator observations and estimated selection through female fitness on flowering phenology and floral traits between two Primula secundiflora populations. We quantified pollinator-mediated selection by subtracting estimates of selection gradients of plants receiving supplemental hand pollination from those of plants receiving open pollination. There was net directional selection for an earlier flowering start date at populations where the dominant pollinators were syrphid flies, and flowering phenology was also subjected to stabilized quadratic selection. However, a later flowering start date was significantly selected at populations where the dominant pollinators were legitimate (normal pollination through the corolla tube entrance) and illegitimate bumblebees (abnormal pollination through nectar robbing hole which located at the corolla tube), and flowering phenology was subjected to disruptive quadratic selection. Wider corolla tube entrance diameter was selected at both populations. Furthermore, the strength of net directional selection on flowering start date and corolla tube entrance diameter was stronger at the population where the dominant pollinators were syrphid flies. Pollinator-mediated selection explained most of the between-population variations in the net directional selection on flowering phenology and corolla tube entrance diameter. Our results suggested the important influence of pollinator-mediated selection on floral evolution. Variations in pollinator assemblages not only resulted in variation in the direction of selection but also the strength of selection on floral traits.

Self- and intra-morph incompatibility and selection analysis of an inconspicuous distylous herb growing on the Tibetan plateau (Primula tibetica).

There is discussion over whether pollen limitation exerts selection on floral traits to increase floral display or selects for traits that promote autonomous self-fertilization. Some studies have indicated that pollen limitation does not mediate selection on traits associated with either pollinator attraction or self-fertilization. Primula tibetica is an inconspicuous cross-fertilized plant that may suffer from pollen limitation. We conducted a selection analysis on P. tibetica to investigate whether pollen limitation results in selection for an increased floral display in case the evolution of autonomous self-fertilization has been difficult for this plant. The self- and intra-morph incompatibility features, the capacity for autonomous self-fertilization, and the magnitude of pollen limitation were examined through hand-pollination experiments. In 2016, we applied selection analysis on the flowering time, corolla width, stalk height, flower tube length, and flower number in P. tibetica by tagging 76 open-pollinated plants and 37 hand-pollinated plants in the field. Our results demonstrated that P. tibetica was strictly self- and intra-morph incompatible. Moreover, the study population underwent severe pollen limitation during the 2016 flowering season. The selection gradients were found to be significantly positive for flowering time, flower number, and corolla width, and marginally significant for the stalk height. Pollinator-mediated selection was found to be significant on the flower number and corolla width, and marginally significant on stalk height. Our results indicate that the increased floral display may be a vital strategy for small distylous species that have faced difficulty in evolving autonomous self-fertilization.

Ginger and the beetle: Evidence of primitive pollination system in a Himalayan endemic alpine ginger (Roscoea alpina, Zingiberaceae).

The Himalayan endemic alpine genus Roscoea, like other members of ginger family, exhibits the combination of floral traits that would fit pollination by long distant foragers such as bees, birds or flies. We studied the pollination biology of Roscoea alpina, observed potential floral visitors and determined their foraging behaviour, visitation frequency and pollination efficiency, to seek evidence in support of the pollination syndrome hypothesis. We also measured the floral spectra of R. alpina flowers to evaluate if signals fit with the currently known framework for observed floral visitors. We found that R. alpina have autonomous selfing and pollinator-mediated crossing, but lack apomixis. We observed that a beetle (Mylabris sp.), and a moth (Macroglossum nycteris) visit the flowers of R. alpina for pollen and nectar feeding respectively. Our field observations, the stigmatic pollen count and fruit set data indicated that the visit by the beetle was legitimate, while that of the moth was illegitimate. Emasculated flowers visited by beetles set as many fruits and seeds/fruit as auto-selfed and naturally pollinated flowers, while emasculated flowers excluded from beetle visits did not set fruit and seed; indicating that a single visit of a beetle to the flowers of R. alpina can facilitate pollination. We found that flower spectral signal of R. alpina does not fit typical spectra previously reported for beetle or bee-visited flowers. Our results suggest that, to ensure reproductive success in alpine habitat, R. alpina has evolved autonomous selfing as a predominant mode of reproduction, while beetle pollination would promote genetic diversity of this plant species. The visitation of beetles to the flowers of R. alpina, despite floral signal mismatch with the classically associated beetle vision, suggests that a different visual processing may operate in this plant-pollinator interaction at high altitudes.

Delivery System Targeting Hemagglutinin of Influenza Virus A to Facilitate Antisense-Based Anti-H1N1 Therapy.

Antisense oligonucleotides (ODNs) are therapeutic molecules that hybridize to complementary target mRNA sequences. To further overcome the poor cellular uptake of ODNs, we proposed a novel strategy to deliver ODNs by conjugating the anti-influenza A virus (IAV) ODN with a peptide showing high affinity to the hemagglutinin (HA) on the surface of IAV particles or the IAV-infected host cells. The HA-specific binding peptides were selected by phage display, and the individual binding clones are characterized by DNA sequencing, and the selected phage was further assayed by enzyme-linked immunosorbent assay. The final selected HA-binding peptide, SHGRITFAYFAN, was conjugated to an anti-IAV ODN. The delivery efficiency and the anti-IAV effects of the conjugated molecule were evaluated in a cell-culture and a mouse-infection model. The conjugated molecule was successfully delivered into IAV-infected host cells more efficiently than the anti-IAV ODN in vitro and in vivo. Furthermore, the conjugated molecule protected 80% of the mice from lethal challenge and inhibited the plaque count by 75% compared to the unconjugated molecule (60% and 40%). These findings demonstrate that the delivery of antisense oligodeoxynucleotides to infected tissues by a virus-binding peptide-mediated system is a potential therapeutic strategy against IAV.

Association between high levels of Notch3 expression and high invasion and poor overall survival rates in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a commonly fatal tumour. It is characterized by early metastasis and high mortality. Many patients die as a result of PDAC tumour progression. However, the underlying mechanism of invasion and metastasis in PDAC is still not fully understood. Previous studies showed that the Notch signalling pathway may play an important role in the progression of tumour invasion and metastasis. However, it is not yet known whether the Notch signalling pathway participates in the progression of invasion in PDAC. In the present study, immunohistochemistry showed that a high expression of Notch3 was correlated with tumour grade, metastasis, venous invasion and American Joint Committee on Cancer (AJCC) tumor-node-metastasis (TNM) stage. Kaplan-Meier curves suggested that a high expression of Notch3 was a significant risk factor for shortened survival time. We also showed that inhibition of Notch3 had an anti­invasion role in PDAC cells. In vitro, the inhibition of Notch3 reduced the migration and invasion capabilities of PDAC cells by regulating the expressions of E-cadherin, CD44v6, MMP-2, MMP-9, VEGF and uPA via regulating the COX-2 and ERK1/2 pathways. These results indicated that downregulation of the Notch signalling pathway may be a novel and useful approach for preventing and treating PDAC invasion.

Protective effects of phillyrin against influenza A virus in vivo.

Influenza A virus infection represents a great threat to public health. However, owing to side effects and the emergence of resistant virus strains, the use of currently available anti-influenza drugs may be limited. In order to identify novel anti-influenza drugs, we investigated the antiviral effects of phillyrin against influenza A virus infection in vivo. The mean survival time, lung index, viral titers, influenza hemagglutinin (HA) protein and serum cytokines levels, and histopathological changes in lung tissue were examined. Administration of phillyrin at a dose of 20 mg/kg/day for 3 days significantly prolonged the mean survival time, reduced the lung index, decreased the virus titers and interleukin-6 levels, reduced the expression of HA, and attenuated lung tissue damage in mice infected with influenza A virus. Taken together, these data showed that phillyrin had potential protective effects against infection caused by influenza A virus.