Advancing screening tool for hospice needs and end-of-life decision-making process in the emergency department.

BACKGROUND: Predicting mortality in the emergency department (ED) is imperative to guide palliative care and end-of-life decisions. However, the clinical usefulness of utilizing the existing screening tools still leaves something to be desired. METHODS: We advanced the screening tool with the A-qCPR (Age, qSOFA (quick sepsis-related organ failure assessment), cancer, Performance Status Scale, and DNR (Do-Not-Resuscitate) risk score model for predicting one-year mortality in the emergency department of Taipei City Hospital of Taiwan with the potential of hospice need and evaluated its performance compared with the existing screening model. We adopted a large retrospective cohort in conjunction with in-time (the trained and the holdout validation cohort) for the development of the A-qCPR model and out-of-time validation sample for external validation and model robustness to variation with the calendar year. RESULTS: A total of 10,474 patients were enrolled in the training cohort and 33,182 patients for external validation. Significant risk scores included age (0.05 per year), qSOFA ≥ 2 (4), Cancer (5), Eastern Cooperative Oncology Group (ECOG) Performance Status score ≥ 2 (2), and DNR status (2). One-year mortality rates were 13.6% for low (score ≦ 3 points), 29.9% for medium (3 < Score ≦ 9 points), and 47.1% for high categories (Score > 9 points). The AUROC curve for the in-time validation sample was 0.76 (0.74-0.78). However, the corresponding figure was slightly shrunk to 0.69 (0.69-0.70) based on out-of-time validation. The accuracy with our newly developed A-qCPR model was better than those existing tools including 0.57 (0.56-0.57) by using SQ (surprise question), 0.54 (0.54-0.54) by using qSOFA, and 0.59 (0.59-0.59) by using ECOG performance status score. Applying the A-qCPR model to emergency departments since 2017 has led to a year-on-year increase in the proportion of patients or their families signing DNR documents, which had not been affected by the COVID-19 pandemic. CONCLUSIONS: The A-qCPR model is not only effective in predicting one-year mortality but also in identifying hospice needs. Advancing the screening tool that has been widely used for hospice in various scenarios is particularly helpful for facilitating the end-of-life decision-making process in the ED.

Acidaminococcus homini s sp. nov., Amedibacillus hominis sp. nov., Lientehia hominis gen. nov. sp. nov., Merdimmobilis hominis gen. nov. sp. nov., and Paraeggerthella hominis sp. nov., isolated from human faeces.

The human gastrointestinal tract is inhabited by various microorganisms, including thousands of bacterial taxa that have yet to be cultured and characterized. In this report, we describe the isolation, cultivation, genotypic and phenotypic characterization and taxonomy of five novel anaerobic bacterial strains that were recovered during the massive cultivation and isolation of gut microbes from human faecal samples. On the basis of the polyphasic taxonomic results, we propose two novel genera and five novel species. They are Acidaminococcus hominis sp. nov. (type strain NSJ-142T=CGMCC 1.17903T=KCTC 25346T), Amedibacillus hominis sp. nov. (type strain NSJ-176T=CGMCC 1.17933T=KCTC 25355T), Lientehia hominis gen. nov. sp. nov. (type strain NSJ-141T=CGMCC 1.17902T=KCTC 25345T), Merdimmobilis hominis gen. nov. sp. nov. (type strain NSJ-153T=CGMCC 1.17915T=KCTC 25350T) and Paraeggerthella hominis sp. nov. (type strain NSJ-152T=CGMCC 1.17914T=KCTC 25349T).

Endocytosis of Peptidase Inhibitor SerpinE2 promotes Myocardial Fibrosis through activating ERK1/2 and ß-catenin Signaling Pathways.

Cardiac fibrosis is one of the common pathological processes in many cardiovascular diseases characterized by excessive extracellular matrix deposition. SerpinE2 is a kind of protein that inhibits peptidase in extracellular matrix and up-regulated tremendously in mouse model of cardiac fibrosis induced by pressure-overloaded via transverse aortic constriction (TAC) surgery. However, its effect on cardiac fibroblasts (CFs), collagen secretion and the underlying mechanism remains unclear. In this study, DyLight® 488 green fluorescent dye or His-tagged proteins were used to label the exogenous serpinE2 protein. It was showed that extracellular serpinE2 translocated into CFs by low-density lipoprotein receptor-related protein 1 (LRP1) and urokinase plasminogen activator receptor (uPAR) of cell membrane through endocytosis. Knockdown of LRP1 or uPAR reduced the level of serpinE2 in CFs and down-regulated the collagen expression. Inhibition of the endocytosis of serpinE2 could inhibit ERK1/2 and ß-catenin signaling pathways and subsequently attenuated collagen secretion. Knockdown of serpinE2 attenuates cardiac fibrosis in TAC mouse. We conclude that serpinE2 could be translocated into cardiac fibroblasts due to endocytosis through directly interact with the membrane protein LRP1 and uPAR, and this process activated the ERK1/2, ß-catenin signaling pathways, consequently promoting collagen production.

Alkaliphilus flagellatus sp. nov., Butyricicoccus intestinisimiae sp. nov., Clostridium mobile sp. nov., Clostridium simiarum sp. nov., Dysosmobacter acutus sp. nov., Paenibacillus brevis sp. nov., Peptoniphilus ovalis sp. nov. and Tissierella simiarum sp. nov., isolated from monkey faeces.

Non-human primates harbour diverse microbiomes in their guts. As a part of the China Microbiome Initiatives, we cultivated and characterized the gut microbiome of cynomolgus monkeys (Macaca fascicularis). In this report, we communicate the characterization and taxonomy of eight bacterial strains that were obtained from faecal samples of captive cynomolgus monkeys. The results revealed that they represented eight novel bacterial species. The proposed names of the eight novel species are Alkaliphilus flagellatus (type strain MSJ-5T=CGMCC 1.45007T=KCTC 15974T), Butyricicoccus intestinisimiae MSJd-7T (MSJd-7T=CGMCC 1.45013T=KCTC 25112T), Clostridium mobile (MSJ-11T=CGMCC 1.45009T=KCTC 25065T), Clostridium simiarum (MSJ-4T=CGMCC 1.45006T=KCTC 15975T), Dysosmobacter acutus (MSJ-2T=CGMCC 1.32896T=KCTC 15976T), Paenibacillus brevis MSJ-6T (MSJ-6T=CGMCC 1.45008T=KCTC 15973T), Peptoniphilus ovalis (MSJ-1T=CGMCC 1.31770T=KCTC 15977T) and Tissierella simiarum (MSJ-40T=CGMCC 1.45012T=KCTC 25071T).

Laminaria japonica Polysaccharide Suppresses Atherosclerosis via Regulating Autophagy-Mediated Macrophage Polarization.

The present work aimed to explore the effect and underlying mechanism of a homogeneous Laminaria japonica polysaccharide (LJP61A) on macrophage polarization in high-fat-diet-fed LDLr-/- mice and Ox-LDL-induced macrophages. Results showed that LJP61A remarkably reduced the lesion burden in atherosclerotic mice, alleviated lipid deposition in Ox-LDL-stimulated macrophages, decreased the expression of M1 macrophage markers, and increased the expression of M2 macrophage markers, thus reducing the M1/M2 macrophage phenotype ratio. Meanwhile, the autophagic flux of macrophages was enhanced by LJP61A treatment in vitro and in vivo. 3-Methyladenine is an autophagic inhibitor. As expected, this inhibitor blocked the effects of LJP61A on macrophage polarization. SIRT1 and FoxO1 are two key upstream genes that control the autophagy behavior. We also found that LJP61A significantly up-regulated the expression of SIRT1 and FoxO1. However, these effects of LJP61A were abolished by the SIRT1 siRNA and FoxO1 inhibitor AS1842856. These results suggested that LJP61A reduced atherosclerosis in HFD-induced LDLr-/- mice via regulating autophagy-mediated macrophage polarization.

LncRNA GAS5 inhibits miR-579-3p to activate SIRT1/PGC-1α/Nrf2 signaling pathway to reduce cell pyroptosis in sepsis-associated renal injury.

Sepsis is a life-threatening condition that can lead to several organ failures including kidney. In this study, we investigated the roles of GAS5 and miR-579-3p in regulating cell pyroptosis in the sepsis-induced renal injury model. Lipopolysaccharide (LPS) treatment or cecal ligation and puncture (CLP) surgery was used to create the in vitro and in vivo sepsis-induced renal injury model. The interactions between GAS5 and miR-579-3p, and miR-579-3p and SIRT1 were determined by bioinformatic prediction, luciferase reporter assay, and RIP assay. In vitro cell pyroptosis was examined by flow cytometry marked with active caspase-1 and PI. The protein levels of IL-1ß and IL-18 induced by cell pyroptosis were quantified using ELISA assay. In vivo renal injuries were evaluated with HE and TUNEL stainings, bacterial load in serum and creatinine, and blood urea nitrogen content analyses. Expression levels of GAS5, miR-579-3p, pyroptosis, and SIRT1/PGC-1a/Nrf2 pathway-related molecules were evaluated by qRT-PCR or Western blot. GAS5 and SIRT1 were downregulated, whereas miR-579-3p was upregulated in in vitro and in vivo sepsis-induced renal injury models. GAS5 negatively and directly regulated miR-579-3p to reduce cell pyroptosis via the activation of SIRT1/PGC-1a/Nrf2 pathway. In addition, miR-579-3p suppressed PGC-1a/Nrf2 pathway to induce cell pyroptosis by directly targeting SIRT1. What's more, overexpression of GAS5, or knockdown of miR-579-3p, enhanced SIRT1 expression that led to the improved survival rate, reduced the weight loss, and relieved renal injuries in septic mice. Overexpression of GAS5 demonstrated protective effects against sepsis-induced renal injury via downregulating miR-579-3p and activating SIRT1/PGC-1α/Nrf2 pathway to inhibit cell pyroptosis.

Meta-analysis of arterial anastomosis techniques in head and neck free tissue transfer.

The present meta-analysis aimed to investigate the differences in the incidence of thrombosis and vascular compromise in arterial anastomosis between microvascular anastomotic devices and hand-sewn techniques during free tissue transfer in the head and neck. We searched for articles in PubMed/Medline, CNKI, WANFANG DATA, Cochrane Central Register of Controlled Trials, Cochrane Database of Systematic Reviews, and Web of Science, from January 1, 1962 till April 1, 2020 that reported data of microvascular anastomosis during free tissue transfer in the head and neck. The incidence of arterial thrombosis or vascular compromise, or both was the primary outcome. The secondary outcome was anastomotic time. We also assessed the sensitivity and the risk of bias. This meta-analysis included 583 arterial anastomoses from six studies. The group using microvascular anastomotic devices tended to have an increased incidence of arterial thrombosis and vascular compromise (risk ratio (RR), 3.42; P = 0.38; 95% confidence interval (CI), 0.91-12.77). The hand-sewn technique took significantly longer to perform the anastomosis compared with that of the microvascular anastomotic devices (weighted mean difference, 15.26 min; P<0.01; 95% CI, 14.65-15.87). Microvascular anastomotic devices might increase the risk of arterial thrombosis and vascular compromise compared with the hand-sewn technique; however, further randomized controlled trials are needed to provide a more accurate estimate. The application of microvascular anastomotic devices will help to reduce anastomotic surgery time and achieve acceptable vessel opening, benefiting from the developments of arterial couplers and microsurgical techniques.

The interaction between C/EBPß and TFAM promotes acute kidney injury via regulating NLRP3 inflammasome-mediated pyroptosis.

Sepsis-induced inflammatory damage is a crucial cause of acute kidney injury (AKI), and AKI is an ecumenical fearful complication in approximately half of patients with sepsis. CCAAT/enhancer-binding protein ß (C/EBPß) plays roles in regulating acute phase responses and inflammation. However, the role and mechanism of C/EBPß in AKI are unclear. LPS combined with ATP-treated renal epithelial cells HK2 and cecal ligation-peferation (CLP)-mice were used as models of AKI in vitro and in vivo. Cell damage, the secretion of interleukin-1 beta (IL-1ß), IL-18 and cysteinyl aspartate specific proteinase 1 (caspase-1) activity were tested by LDH, ELISA assay and flow cytometry analysis, respectively. The expression levels of TFAM, C/EBPß, and pyroptosis-related molecules were tested by qRT-PCR and Western blotting. Chromatin immunoprecipitation (ChIP) assessed the interaction between C/EBPß with TFAM. Hematoxylin-Eosin (H&E) staining detected pathological changes of kidney tissues, and immunohistochemistry measured TFAM and C/EBPß in mice kidney tissues. C/EBPß or TFAM were up-regulated in LPS combined with ATP -induced HK2 cells. Knockdown of C/EBPß could suppress cell injury and the secretion of IL-1ß and IL-18 induced by LPS combined with ATP. Furthermore, C/EBPß up-regulated the expression levels of TFAM via directly binding to TFAM promoter. Overexpression of TFAM reversed the effects of C/EBPß deficiency on pyroptosis. Knockdown of C/EBPß could inhibit NLRP3 inflammasome-mediated caspase-1 signaling pathway by inactivating TFAM/RAGE pathway. It was further confirmed in the AKI mice that C/EBPß and TFAM promoted AKI by activating NLRP3-mediated pyroptosis. The interaction of between C/EBPß and TFAM facilitated pyroptosis by activating NLRP3/caspase-1 signal axis, thereby promoting the occurrence of AKI.

Upregulation of Mitochondrial Transcription Factor A Promotes the Repairment of Renal Tubular Epithelial Cells in Sepsis by Inhibiting Reactive Oxygen Species-Mediated Toll-Like Receptor 4/p38MAPK Signaling.

BACKGROUND: Mitochondrial transcription factor A (TFAM) plays multiple pathophysiologic roles in mitochondrial DNA (mtDNA) maintenance. However, the role of TFAM in sepsis-induced acute kidney injury (AKI) remains largely unknown. METHODS: Lipopolysaccharide (LPS) treatment of HK-2 cells mimics the in vitro model of AKI inflammation. pcDNA3.1 plasmid was used to construct pcDNA3.1-TFAM. sh-TFAM-543, sh-TFAM-717, sh-TFAM-765, sh-TFAM-904 and pcDNA3.1-TFAM were transfected into HK-2 cells using Lipofectamine 2000. MtDNA transcriptional levels were detected by quantitative real-time polymerase chain reaction (qRT-PCR). 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay was performed to assess the cell viability. Changes in reactive oxygen species (ROS) and mitochondrial membrane potential in HK-2 cells were detected using the corresponding kits. Immunofluorescence experiment was used to investigate the displacement of TFAM. mRNA and protein expression levels of TFAM and its related genes were measured by qRT-PCR and western blot respectively. Mice in sepsis were administered cecal ligation and puncture surgery. RESULTS: LPS treatment was a non-lethal influencing factor, leading to the upregulation of ROS levels and downregulation of mtDNA copy number and NADH dehydrogenase subunit-1 (ND1) expression, and caused damage to the mitochondria. As the LPS treatment time increased, TFAM was displaced from the periphery of the nucleus to cytoplasm. TFAM reduced ROS and P38MAPK levels by inhibiting toll-like receptor 4 (TLR4) expression, ultimately inhibiting inflammation and repairing mtDNA. CONCLUSIONS: Our results indicate that TFAM repairs mtDNA by blocking the TLR4/ROS/P38MAPK signaling pathway in inflammatory cells, thereby repairing septic tubular epithelial cells, and TFAM may serve as a new target for sepsis therapy.

Parabacteroides acidifaciens sp. nov., isolated from human faeces.

A polyphasic taxonomic approach was applied to characterize an anaerobic bacterial strain, 426-9T, that was isolated from human faeces. The strain was Gram-stain-negative, non-motile, non-spore-forming, non-pigmented and rod-shaped. Strain 426-9T grew anaerobically at 20-45 °C (optimally at 37-40 °C) and at pH 6.0-10.0 (optimally at pH 6.0-8.0). The major polar lipids were phosphatidylethanolamine, seven amino phospholipids and three phospholipids. The major fatty acids of strain 426-9T were anteiso-C15 : 0 and iso-C17 : 0 3-OH, and the predominant respiratory quinones were menaquinones MK-9 and MK-10. End-products of glucose fermentation were acetate, propionate, iso-butyrate and iso-pentanoate. 16S rRNA gene sequence analysis showed that strain 426-9T was a member of the genus Parabacteroides. The level of 16S rRNA gene sequence similarity of strain 426-9T to the type species of the genus, Parabacteroides distasonis ATCC 8503T, was 91.0 %. Within the genus Parabacteroides, strain 426-9T was phylogenetically closely related to Parabacteroides johnsonii M-165T (96.0 % 16S rRNA gene sequence similarity). The draft genome of strain 426-9T comprised 5.15 Mb with a DNA G+C content of 45.9 mol%. A total of 4088 genes were predicted and, of those, 3744 were annotated. On the basis of phenotypic, chemotaxonomic and phylogenetic characterization, strain 426-9T represents a novel species within the genus Parabacteroides, for which the name Parabacteroides acidifaciens sp. nov. is proposed. The type strain is 426-9T (=CGMCC 1.13558T=NBRC 113433T).

Effects of Catalytic Action and Ligand Binding on Conformational Ensembles of Adenylate Kinase.

Crystal structures of adenylate kinase (AdK) from Escherichia coli capture two states: an "open" conformation (apo) obtained in the absence of ligands and a "closed" conformation in which ligands are bound. Other AdK crystal structures suggest intermediate conformations that may lie on the transition pathway between these two states. To characterize the transition from open to closed states in solution, X-ray solution scattering data were collected from AdK in the apo form and with progressively increasing concentrations of five different ligands. Scattering data from apo AdK are consistent with scattering predicted from the crystal structure of AdK in the open conformation. In contrast, data from AdK samples saturated with Ap5A do not agree with that calculated from AdK in the closed conformation. Using cluster analysis of available structures, we selected representative structures in five conformational states: open, partially open, intermediate, partially closed, and closed. We used these structures to estimate the relative abundances of these states for each experimental condition. X-ray solution scattering data obtained from AdK with AMP are dominated by scattering from AdK in the open conformation. For AdK in the presence of high concentrations of ATP and ADP, the conformational ensemble shifts to a mixture of partially open and closed states. Even when AdK is saturated with Ap5A, a significant proportion of AdK remains in a partially open conformation. These results are consistent with an induced-fit model in which the transition of AdK from an open state to a closed state is initiated by ATP binding.

Adoptive Regulatory T-cell Therapy Attenuates Perihematomal Inflammation in a Mouse Model of Experimental Intracerebral Hemorrhage.

The CD4+CD25+ regulatory T cells (Tregs), an innate immunomodulator, suppress cerebral inflammation and maintain immune homeostasis in multiple central nervous system injury, but its role in intracerebral hemorrhage (ICH) has not been fully characterized. This study investigated the effect of Tregs on brain injury using the mouse ICH model, which is established by autologous blood infusion. The results showed that tail intravenous injection of Tregs significantly reduced brain water content and Evans blue dye extravasation of perihematoma at day (1, 3 and 7), and improved short- and long-term neurological deficits following ICH in mouse model. Tregs treatment reduced the content of pro-inflammatory cytokines interleukin (IL)-1ß, IL-6, tumor necrosis factor-α, and malondialdehyde, while increasing the superoxide dismutase (SOD) enzymatic activity at day (1, 3 and 7) following ICH. Furthermore, Tregs treatment obviously reduced the number of NF-κB+, IL-6+, TUNEL+ and active caspase-3+ cells at day 3 after ICH. These results indicate that adoptive transfer of Tregs may provide neuroprotection following ICH in mouse models.

Lactobacillus rhamnosus GG attenuates interferon-{gamma} and tumour necrosis factor-alpha-induced barrier dysfunction and pro-inflammatory signalling.

The intestinal epithelium forms a protective barrier against luminal contents and the external environment, mediated via intercellular tight junctions (TJs). The TJ can be disrupted via cell signalling induced by either enteric pathogens or pro-inflammatory cytokines, thereby contributing to various intestinal disorders ranging from acute infectious diarrhoea to chronic inflammatory bowel diseases. Probiotics, such as Lactobacillus rhamnosus GG (LGG), are reported to confer beneficial effects on epithelial cells, including antagonizing infections and reducing overt pro-inflammatory responses, but the underlying mechanisms of these observed effects require further characterization. We hypothesized that probiotics preserve barrier function by interfering with pro-inflammatory cytokine signalling. Caco-2bbe cells were seeded into Transwells to attain polarized monolayers with intercellular TJs. Monolayers were inoculated apically with the probiotic LGG 3 h prior to the addition of IFN-γ (100 ng ml(-1)) to the basolateral medium overnight. The monolayers were then placed in fresh basal medium±TNF-α (10 ng ml(-1)) and transepithelial electrical resistance (TER) measurements were taken over the time-course of TNF-α stimulation. To complement the TER findings, cells were processed for zona occludens-1 (ZO-1) immunofluorescence staining. As a measure of TNF-α downstream signalling, cells were immunofluorescently stained for NF-κB p65 subunit and CXCL-8 mRNA was quantified by qRT-PCR. Basal cell culture medium was collected after overnight TNF-α stimulation to measure secreted chemokines, including CXCL-8 (interleukin-8) and CCL-11 (eotaxin). Following LGG inoculation, IFN-γ priming and 24 h TNF-α stimulation, epithelial cells maintained TER and ZO-1 distribution. LGG diminished the nuclear translocation of p65, demonstrated by both immunofluorescence and CXCL-8 mRNA expression. CXCL-8 and CCL-11 protein levels were decreased in LGG-inoculated, cytokine-challenged cells. These findings indicate that LGG alleviates the effects of pro-inflammatory cytokines on epithelial barrier integrity and inflammation, mediated, at least in part, through inhibition of NF-κB signalling.

[Expression of the proteins associated with transforming growth factor-beta/Smad signaling pathway in Peutz-Jeghers syndrome].

OBJECTIVE: To detect the expression of important proteins associated with transforming growth factor-beta (TGF-beta)/Smad signaling pathway in Peutz-Jeghers syndrome (PJS) and investigate the correlation of these proteins to LKB1 gene expression. METHODS: The expression and localization of LKB1, TGFbeta1 and pSmad2 proteins in 20 PJS polyp samples and normal intestinal mucosal tissues were detected with immunohistochemical staining. RESULTS: The expressions of LKB1, TGFbeta1 and pSmad2 were lower in PJS polyps than in normal mucosa, and the differences in LKB1 and TGFbeta1 proteins were significantly different between them (P<0.05). In PJS polyps, positive correlations were found between LKB1 and TGFbeta1 and between TGFbeta1 and pSmad2 expressions. CONCLUSION: TGFbeta/Smad pathway is probably subjected to the regulation by LKB1 and may play a role in the occurrence of PJS.

Thermosensitive liposomes entrapping iron oxide nanoparticles for controllable drug release.

Iron oxide nanoparticles can serve as a heating source upon alternative magnetic field (AMF) exposure. Iron oxide nanoparticles can be mixed with thermosensitive nanovehicles for hyperthermia-induced drug release, yet such a design and mechanism may not be suitable for controllable drug release applications in which the tissues are susceptible to environmental temperature change such as brain tissue. In the present study, iron oxide nanoparticles were entrapped inside of thermosensitive liposomes for AMF-induced drug release while the environmental temperature was maintained at a constant level. Carboxyfluorescein was co-entrapped with the iron oxide nanoparticles in the liposomes as a model compound for monitoring drug release and environmental temperature was maintained with a water circulator jacket. These experiments have been successfully performed in solution, in phantom and in anesthetized animals. Furthermore, the thermosensitive liposomes were administered into rat forearm skeletal muscle, and the release of carboxylfluorescein triggered by the external alternative magnetic field was monitored by an implanted microdialysis perfusion probe with an on-line laser-induced fluorescence detector. In the future such a device could be applied to simultaneous magnetic resonance imaging and non-invasive drug release in temperature-sensitive applications.

Radiation and diversification within the Ligularia-Cremanthodium-Parasenecio complex (Asteraceae) triggered by uplift of the Qinghai-Tibetan Plateau.

The Ligularia-Cremanthodium-Parasenecio (L-C-P) complex of the Tussilagininae (Asteraceae: Senecioneae) contains more than 200 species that are endemic to the Qinghai-Tibetan Plateau in eastern Asia. These species are morphologically distinct; however, their relationships appear complex. A phylogenetic analysis of members of the complex and selected taxa of the tribe Senecioneae was conducted using chloroplast (ndhF and trnL-F) and nuclear (ITS) sequences. Phylogenetic trees were constructed from individual and combined datasets of the three different sequences. All analyses suggested that Doronicum, a genus that has been included in the Tussilagininae, should be excluded from this subtribe and placed at the base of the tribe Senecioneae. In addition, the Tussilagininae should be broadly circumscribed to include the Tephroseridinae. Within the expanded Tussilagininae containing all 13 genera occurring in eastern Asia, Tussilago and Petasites diverged early as a separate lineage, while the remaining 11 genera comprise an expanded L-C-P complex clade. We suggest that the L-C-P clade, which is largely unresolved, most likely originated as a consequence of an explosive radiation. The few monophyletic subclades identified in the L-C-P clade with robust support further suggest that some genera of Tussilagininae from eastern Asia require generic re-circumscriptions given the occurrence of subclades containing species of the same genus in different parts of the phylogentic tree due to homoplasy of important morphological characters used to delimit them. Molecular-clock analyses suggest that the explosive radiation of the L-C-P complex occurred mostly within the last 20 million years, which falls well within the period of recent major uplifts of the Qinghai-Tibetan Plateau between the early Miocene to the Pleistocene. It is proposed that significant increases in geological and ecological diversity that accompanied such uplifting, most likely promoted rapid and continuous allopatric speciation in small and isolated populations, and allowed fixation or acquisition of similar morphological characters within unrelated lineages. This phenomenon, possibly combined with interspecific diploid hybridization because of secondary sympatry during relatively stable stages between different uplifts, could be a major cause of high species diversity in the Qinghai-Tibetan Plateau and adjacent areas of eastern Asia.