[Prediction and analysis of Q-markers of Elephantopus scaber based on its UPLC fingerprint, content determination of components, and in vitro a nti-tumor activity].

This study aimed to provide scientific evidence for predicting quality markers(Q-markers) of Elephantopus scaber by establishing UPLC fingerprint of E. scaber from different geographical origins and determining the content of 13 major components, as well as conducting in vitro anti-cancer activity investigation of the main components. The chromatographic column used was Waters CORTECS UPLC C_(18)(2.1 mm×150 mm, 1.6 µm), and the mobile phase consisted of acetonitrile and 0.1% formic acid solution(gradient elution). The column temperature was set at 30 ℃, and the flow rate was 0.2 mL·min~(-1). The injection volume was 1 µL, and the detection wavelength was 240 nm. The UPLC fingerprint of E. scaber was fitted using the Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine(2012 edition) to determine common peaks, evaluate similarity, identify and determine the content of major components. The CCK-8 assay was used to explore the inhibitory effect of the main components on the proliferation of lung cancer cells. The results showed that in the established UPLC fingerprint of E. scaber, 35 common peaks were identified. Thirteen major components, including neochlorogenic acid(peak 1), chlorogenic acid(peak 2), cryptochlorogenic acid(peak 3), caffeic acid(peak 4), schaftoside(peak 6), galuteolin(peak 9), isochlorogenic acid B(peak 10), isochlorogenic acid A(peak 12), isochlorogenic acid C(peak 18), deoxyelephantopin(peak 28), isodeoxyelephantopin(peak 29), isoscabertopin(peak 31), and scabertopin(peak 32) were identified and quantified, and a quantitative analysis method was established. The results of the in vitro anti-cancer activity study showed that deoxyelephantopin, isodeoxyelephantopin, isoscabertopin, and scabertopin in E. scaber exhibited inhibition rates of lung cancer cell proliferation exceeding 80% at a concentration of 10 µmol·L~(-1), higher than the positive drug paclitaxel. These results indicate that the fingerprint of E. scaber is highly characteristic, and the quantitative analysis method is accurate and stable, providing references for the research on quality standards of E. scaber. Four sesquiterpene lactones in E. scaber show significant anti-cancer activity and can serve as Q-markers for E. scaber.

Diterbutyl phthalate attenuates osteoarthritis in ACLT mice via suppressing ERK/c-fos/NFATc1 pathway, and subsequently inhibiting subchondral osteoclast fusion.

Osteoarthritis (OA) is the most common arthritis with a rapidly increasing prevalence. Disease progression is irreversible, and there is no curative therapy available. During OA onset, abnormal mechanical loading leads to excessive osteoclastogenesis and bone resorption in subchondral bone, causing a rapid subchondral bone turnover, cyst formation, sclerosis, and finally, articular cartilage degeneration. Moreover, osteoclast-mediated angiogenesis and sensory innervation in subchondral bone result in abnormal vascularization and OA pain. The traditional Chinese medicine Panax notoginseng (PN; Sanqi) has long been used in treatment of bone diseases including osteoporosis, bone fracture, and OA. In this study we established two-dimensional/bone marrow mononuclear cell/cell membrane chromatography/time of flight mass spectrometry (2D/BMMC/CMC/TOFMS) technique and discovered that diterbutyl phthalate (DP) was the active constituent in PN inhibiting osteoclastogenesis. Then we explored the therapeutic effect of DP in an OA mouse model with anterior cruciate ligament transaction (ACLT). After ACLT was conducted, the mice received DP (5 mg·kg-1·d-1, ip) for 8 weeks. Whole knee joint tissues of the right limb were harvested at weeks 2, 4, and 8 for analysis. We showed that DP administration impeded overactivated osteoclastogenesis in subchondral bone and ameliorated articular cartilage deterioration. DP administration blunted aberrant H-type vessel formation in subchondral bone marrow and alleviated OA pain assessed in Von Frey test and thermal plantar test. In RANKL-induced RAW264.7 cells in vitro, DP (20 µM) retarded osteoclastogenesis by suppressing osteoclast fusion through inhibition of the ERK/c-fos/NFATc1 pathway. DP treatment also downregulated the expression of dendritic cell-specific transmembrane protein (DC-STAMP) and d2 isoform of the vacuolar (H+) ATPase V0 domain (Atp6v0d2) in the cells. In conclusion, we demonstrate that DP prevents OA progression by inhibiting abnormal osteoclastogenesis and associated angiogenesis and neurogenesis in subchondral bone.

[Advanced progress of main pharmacology activities of triptolide].

Triptolide is an active component in traditional Chinese medicine Tripterygium wilfordii. Currently, triptolide has been used to treat various diseases, including lupus, cancer, rheumatoid arthritis and nephritic syndrome. Its main pharmacology efficacies include anti-inflammation, anti-tumor, and immunity suppression. Recent studies have also demonstrated triptolide's protective effect on cardiovascular diseases and osteoporosis. This paper summarizes the pharmacological efficacy of triptolide based on the advance in studies of triptolide.

LRP1 facilitates hepatic glycogenesis by improving the insulin signaling pathway in HFD-fed mice.

BACKGROUND: LDL receptor-related protein-1 (LRP1) is a cell-surface receptor that functions in diverse physiological pathways. We previously demonstrated that hepatocyte-specific LRP1 deficiency (hLRP1KO) promotes diet-induced insulin resistance and increases hepatic gluconeogenesis in mice. However, it remains unclear whether LRP1 regulates hepatic glycogenesis. METHODS: Insulin signaling, glycogenic gene expression, and glycogen content were assessed in mice and HepG2 cells. The pcDNA 3.1 plasmid and adeno-associated virus serotype 8 vector (AAV8) were used to overexpress the truncated ß-chain (ß∆) of LRP1 both in vitro and in vivo. RESULTS: On a normal chow diet, hLRP1KO mice exhibited impaired insulin signaling and decreased glycogen content. Moreover, LRP1 expression in HepG2 cells was significantly repressed by palmitate in a dose- and time-dependent manner. Both LRP1 knockdown and palmitate treatment led to reduced phosphorylation of Akt and GSK3ß, increased levels of phosphorylated glycogen synthase (GYS), and diminished glycogen synthesis in insulin-stimulated HepG2 cells, which was restored by exogenous expression of the ß∆-chain. By contrast, AAV8-mediated hepatic ß∆-chain overexpression significantly improved the insulin signaling pathway, thus activating glycogenesis and enhancing glycogen storage in the livers of high-fat diet (HFD)-fed mice. CONCLUSION: Our data revealed that LRP1, especially its ß-chain, facilitates hepatic glycogenesis by improving the insulin signaling pathway, suggesting a new therapeutic strategy for hepatic insulin resistance-related diseases.

[Expert consensus on integrated traditional Chinese and western medicine diagnosis and treatment for osteoporotic fractures].

Osteoporotic fractures represent the most severe complications of osteoporosis,characterized by insidious onset,high mortality and disability rates,and a steadily increasing incidence,imposing a significant socioeconomic burden. Western medicine has advantages in diagnosis and surgical interventions,while traditional Chinese medicine excels in holistic management and the restoration of bodily equilibrium. The integration of both traditional Chinese medicine (TCM) and western medicine emerges as an effective therapeutic strategy for osteoporotic fractures. In order to propagate the concept of integrated diagnosis and treatment,foster the advancement of integrated medical techniques for osteoporotic fractures,and establish standardized and normative protocols for disease prevention,diagnosis,and treatment,a consensus expert group,led by Geriatric Branch of Chinese Geriatrics Society,the Young Osteoporosis Group of Orthopedics Branch of Chinese Medical Association,Osteoporosis Group of Orthopedics Branch of Chinese Physician Association,and Osteoporosis Professional Committee of the Shanghai Society of Integrated Traditional Chinese and Western Medicine,was established. This group engaged in deliberations and formulated the "Expert Consensus on Integrated Traditional Chinese and Western Medicine Diagnosis and Treatment of Osteoporotic Fractures" elucidating the concept of integrated medicine and offering recommendations in the domains of prevention,diagnosis,and treatment,with the aspiration of ameliorating the prognosis of osteoporotic fractures and enhancing the quality of life for these patients.

Diets intervene osteoporosis via gut-bone axis.

Osteoporosis is a systemic skeletal disease that seriously endangers the health of middle-aged and older adults. Recently, with the continuous deepening of research, an increasing number of studies have revealed gut microbiota as a potential target for osteoporosis, and the research concept of the gut-bone axis has gradually emerged. Additionally, the intake of dietary nutrients and the adoption of dietary patterns may affect the gut microbiota, and alterations in the gut microbiota might also influence the metabolic status of the host, thus adjusting bone metabolism. Based on the gut-bone axis, dietary intake can also participate in the modulation of bone metabolism by altering abundance, diversity, and composition of gut microbiota. Herein, combined with emerging literatures and relevant studies, this review is aimed to summarize the impacts of different dietary components and patterns on osteoporosis by acting on gut microbiota, as well as underlying mechanisms and proper dietary recommendations.

Bone-organ axes: bidirectional crosstalk.

In addition to its recognized role in providing structural support, bone plays a crucial role in maintaining the functionality and balance of various organs by secreting specific cytokines (also known as osteokines). This reciprocal influence extends to these organs modulating bone homeostasis and development, although this aspect has yet to be systematically reviewed. This review aims to elucidate this bidirectional crosstalk, with a particular focus on the role of osteokines. Additionally, it presents a unique compilation of evidence highlighting the critical function of extracellular vesicles (EVs) within bone-organ axes for the first time. Moreover, it explores the implications of this crosstalk for designing and implementing bone-on-chips and assembloids, underscoring the importance of comprehending these interactions for advancing physiologically relevant in vitro models. Consequently, this review establishes a robust theoretical foundation for preventing, diagnosing, and treating diseases related to the bone-organ axis from the perspective of cytokines, EVs, hormones, and metabolites.

Targeting the gut microbiota-related metabolites for osteoporosis: The inextricable connection of gut-bone axis.

Osteoporosis is a systemic skeletal disease characterized by decreased bone mass, destruction of bone microstructure, raised bone fragility, and enhanced risk of fractures. The correlation between gut microbiota and bone metabolism has gradually become a widespread research hotspot in recent years, and successive studies have revealed that the alterations of gut microbiota and its-related metabolites are related to the occurrence and progression of osteoporosis. Moreover, several emerging studies on the relationship between gut microbiota-related metabolites and bone metabolism are also underway, and extensive research evidence has indicated an inseparable connection between them. Combined with latest literatures and based on inextricable connection of gut-bone axis, this review is aimed to summarize the relation, potential mechanisms, application strategies, clinical application prospects, and existing challenges of gut microbiota and its-related metabolites on osteoporosis, thus updating the knowledge in this research field and providing certain reference for future researches.

Linking the relationship between dietary folic acid intake and risk of osteoporosis among middle-aged and older people: A nationwide population-based study.

Among middle-aged and older people, balanced and nutritious diets are the foundation for maintaining bone health and preventing osteoporosis. This study is aimed at investigating the link between dietary folic acid intake and the risk of osteoporosis among middle-aged and older people. A total of 20,686 people from the National Health and Nutritional Examination Survey (NHANES) 2007-2010 are screened and included, and 5312 people aged ≥45 years with integral data are ultimately enrolled in evaluation. Demographics and dietary intake-related data are gathered and analyzed, and the odds ratio (OR) and 95% confidence interval (CI) of each tertile category of dietary folic acid intake and each unit increase in folic acid are assessed via multivariate logistic regression models. On this basis, the receiver operating characteristic (ROC) curve is used to identify the optimal cutoff value of dietary folic acid intake for indicating the risk of osteoporosis. Of 5312 people with a mean age of 62.4 ± 11.0 years old, a total of 513 people with osteoporosis are screened, and the dietary folic acid intake amount of the osteoporosis group is significantly lower than that of the non-osteoporosis group (p < .001). The lowest tertile category is then used to act as a reference category, and a higher dietary folic acid intake amount is observed to be positively related to lower odds for risk of osteoporosis. This trend is also not changed in adjustments for combinations of different covariates (p all < .05). Based on this, a dietary folic acid intake of 475.5 µg/day is identified as an optimal cutoff value for revealing osteoporosis. Collectively, this nationwide population-based study reveals that a higher daily dietary folic acid intake has potential protective effects on osteoporosis in middle-aged and older people.

The Relationship Between the Effectiveness of HMME-PDT and the Dermoscopic Features of Port-wine Stains in Chinese Pediatric Patients: A Retrospective Study.

INTRODUCTION: Although pulsed dye laser (PDL) remains the gold standard for the treatment of port-wine stains (PWS), hematoporphyrin monomethyl ether photodynamic therapy (HMME-PDT) is another treatment modality that has been shown to be effective in the treatment of PWS. This study aimed to observe the clinical efficacy and therapeutic response of HMME-PDT in the treatment of pediatric Chinese patients with PWS and to analyze the association between the efficacy of therapy and the dermoscopic features of PWS. METHODS: Pediatric patients with PWS and negative HMME skin test were enrolled between December 2017 and May 2021. Patients received an intravenous injection of 5 mg/kg HMME, and lesions were irradiated with 532-nm LED green light with a power density of 70-80 mW/cm2 for 20-25 min. Digital photographs and dermoscopic images were taken before and after two treatment sessions, and the clinical response was observed. The relationship between the efficacy of HMME-PDT and the dermoscopic features of PWS was retrospectively analyzed. RESULTS: A total of 216 pediatric patients (1-14 years) were recruited. Sixty-six patients had the pink type, while 150 had the purple type. After two HMME-PDT sessions, 55 patients showed excellent efficacy (25.46%), 77 patients showed good efficacy (35.65%), 69 patients showed fair efficacy (31.94%), and 15 patients showed no improvement (6.95%). Dotted and globular vessels were highly associated with excellent efficacy (41.82%); linear vessels were mainly associated with good efficacy (54.55%); reticular vessels were mainly associated with fair (55.07%) and mixed vessels were mainly associated with no improvement (26.66%). CONCLUSION: HMME-PDT is an effective and safe treatment for pediatric patients with PWS. Dotted and globular vessels as well as linear vessels showed better efficacy compared to the other dermoscopic patterns in patients with PWS. Dermoscopy can provide useful clinical information about treatment outcomes.

The Efficacy of Hematoporphyrin Monomethyl Ether Photodynamic Therapy in Adult Patients with Port-Wine Stains: A Retrospective Study.

INTRODUCTION: Hematoporphyrin monomethyl ether-photodynamic therapy (HMME-PDT) has been showing promising results in the treatment of port-wine stains (PWSs). We evaluated the clinical efficacy and treatment response of HMME-PDT in adult Chinese patients with PWSs. METHODS: A single-center retrospective study recruited adult PWS patients with negative HMME skin test results from December 2017 to May 2020. Patients received an intravenous injection of 5 mg/kg HMME and the lesions were exposed to 532 nm LED green light with an irradiation power density of 85-95 mW/cm2 for 20-25 min. Digital photographs were taken before and after two therapy sessions and observed by three blinded dermatologists for clinical response. RESULTS: A total of 72 patients aged between 18 and 55 years were recruited. There were 65 patients of the flat purple type, 5 of the hypertrophic type, and 2 of the nodular thickening type. Of the 65 patients, 7 showed excellent efficacy (10.77%), 13 patients indicated good efficacy (20.00%), 47 patients showed fair efficacy (64.62%), while 3 cases displayed no improvement (4.62%). All five patients of the purple and hypertrophic type showed fair efficacy (100%), and no improvement was observed in patients of the nodular thickening type (100%). Pain, pruritus, and a burning sensation were observed during treatment. Edema was noted on the treated areas post-treatment. No other obvious systemic adverse reactions were observed. CONCLUSION: HMME-PDT is an effective and safe treatment for adult patients with purple PWSs. Multiple HMME-PDT treatments can improve the response and cure rate.

Electroacupuncture reduces chronic itch via cannabinoid CB1 receptors in the ventrolateral periaqueductal gray.

Chronic itch severely reduces the quality of life of patients. Electroacupuncture (EA) is widely used to treat chronic itch. However, the underlying mechanism of this therapeutic action of EA is largely unknown. Cannabinoid CB1 receptors in the ventrolateral periaqueductal gray (vlPAG) mediate the analgesic effect of EA. Using a dry skin-induced itch model in mice, we determined whether EA treatment reduces chronic itch via CB1 receptors in the vlPAG. We showed that the optimal inhibitory effect of EA on chronic itch was achieved at the high frequency and high intensity (100 Hz and 3 mA) at "Quchi" (LI11) and "Hegu" (LI14) acupoints, which are located in the same spinal dermatome as the cervical skin lesions. EA reversed the increased expression of CB1 receptors in the vlPAG and decreased the concentration of 5-hydroxytryptamine (5-HT) in the medulla oblongata and the expression of gastrin-releasing peptide receptors (GRPR) in the cervical spinal cord. Furthermore, knockout of CB1 receptors on GABAergic neurons in the vlPAG attenuated scratching behavior and the 5-HT concentration in the medulla oblongata. In contrast, knockout of CB1 receptors on glutamatergic neurons in the vlPAG blocked the antipruritic effects of EA and the inhibitory effect of EA on the 5-HT concentration in the medulla oblongata. Our findings suggest that EA treatment reduces chronic itch by activation of CB1 receptors on glutamatergic neurons and inhibition of CB1 receptors on GABAergic neurons in the vlPAG, thereby inhibiting the 5-HT release from the medulla oblongata to GRPR-expressing neurons in the spinal cord. Our findings suggest that EA attenuates chronic itch via activating CB1 receptors expressed on glutamatergic neurons and downregulating CB1 receptors on GABAergic neurons in the vlPAG, leading to the reduction in 5-HT release in the rostroventral medulla and GRPR signaling in the spinal cord. Our study not only advances our understanding of the mechanisms of the therapeutic effect of EA on chronic itch but also guides the selection of optimal parameters and acupoints of EA for treating chronic itch.

Using side-bending radiographs to determine the distal fusion level in patients with single thoracic idiopathic scoliosis undergoing posterior correction with pedicle screws.

STUDY DESIGN: Prospective. OBJECTIVES: To evaluate a strategy to determine the distal fusion level in posterior pedicle screw correction of single thoracic idiopathic scoliosis. SUMMARY OF BACKGROUND DATA: No standard method for selecting the lowest instrumented vertebra (LIV) for the correction of thoracic adolescent idiopathic scoliosis with posterior all-pedicle screw instrumentations exists. METHODS: Thirty-eight patients with single right thoracic (Lenke 1A) adolescent idiopathic scoliosis undergoing posterior pedicle screw fixation were studied. The LIV was determined using guidelines based on preoperative side-bending radiographs. In brief, (1) the whole thoracic Cobb curve should be included in the fusion mass, and the LIV should not be superior to the lower-end vertebra of the Cobb measurement. (2) On the right side-bending radiographs, the LIV should be derotated to neutral in skeletally immature (Risser 0 to 3) patients and the disc immediately below the LIV must open on the left side by at least 5 degrees. (3) On the left side-bending radiographs, the disc immediately below the LIV must be open on the right side by at least 0 degree. The first segment meeting the criteria when proceeding from the lower-end vertebra caudally is chosen as the LIV. Outcomes were based on the standing radiographs. RESULTS: Minimum follow-up was 2 years. The mean preoperative thoracic curve was 48.4±9.2 degrees and 12.6±6.1 degrees at final follow-up, resulting in a mean correction of 74.7%±8.5%. The mean preoperative compensatory lumbar curve of 23.7±7.5 degrees was 6.3±4.8 degrees at final follow-up. A change in lumbar lordosis from -41.2±11.9 degrees preoperatively to -38.2±9.9 degrees at final follow-up occurred. All patients achieved coronal balance and no decompensation or adding-on phenomenon was observed. Compared with the recommended fusion end by the Harrington stable zone method, 86.9% patients were saved 1 or more motion segment. CONCLUSIONS: The method described was effective in obtaining satisfactory curve correction, adequate trunk balance, and preservation of motion segments.

Comparison between radial head replacement and open reduction and internal fixation in clinical treatment of unstable, multi-fragmented radial head fractures.

The objective of this study was to compare replacement of the radial head by metal prostheses with open reduction and internal fixation (ORIF) for the treatment of unstable, multi-fragmented radial head fractures. A prospective randomised controlled trial was employed to investigate 45 patients with unstable, multi-fragmented fractures of the radial head, from January 2004 to June 2007. The patients were randomised to two groups: the ORIF group and the radial head replacement group. Over the next two years, follow-up assessments recorded Broberg and Morrey scores and postoperative complication rate. Statistical analysis was performed. According to Broberg and Morrey scores, patients receiving radial head replacement achieved significantly better clinical results with 91% (20/22) good or excellent compared to patients assigned to the ORIF group with 65.2% (15/23) good or excellent results (P < 0.01). Postoperative complication rate of the radial head replacement group (13.6%) was significantly lower than that of the ORIF group (47.9%; P < 0.01). Compared with open reduction and internal fixation, radial head replacement with a metal prostheses resulted in favourable joint function for the unstable, multi-fragmented fractures of the radial head.

Shape memory Ni-Ti alloy swan-like bone connector for treatment of humeral shaft nonunion.

From August 1990 to December 2007, 156 patients with humeral shaft nonunion were treated with our patented Ni-Ti shape memory alloy swan-like memory pressure connector (SMC). The SMC device cooled with ice before implantation was warmed to 40-50 degrees C after implantation to produce balanced axial and compression forces to stabilise the fracture three-dimensionally. This combined with autologous bone grafting achieved bone tissue regeneration in the fracture and promoted smooth recovery of joint function, with a nonunion healing rate of 98.7% after a single SMC implantation. Failure of nonunion healing occurred in only two cases but was successfully managed by a further operation. Complications were not found in any of these patients apart from four with pre-existing radial nerve injuries. These results demonstrate the effectiveness of the SMC device for the management of humeral shaft nonunion. The device provides continuous compression of the fracture with minimal trauma to the local blood supply.

Three-dimensional finite element analysis of lumbar vertebra loaded by static stress and its biomechanical significance.

OBJECTIVE: To explore the mechanical behavior of lumbar spine loaded by stress and provide the mechanical basis for clinical analysis and judgement of lumbar spine fracture classification, mechanical distribution and static stress. METHODS: By means of computer simulation method, the constructed lumbar spine three-dimensional model was introduced into three-dimensional finite element analysis by software Ansys 7.0. The lumbar spine mechanical behavior in different parts of the stress loading were calculated. Impact load is 0-8000 N. The peak value was 8000 N. The loading time is 0-40 minutes. The values of the main stress, stress distribution and the lumbar spine unit displacement in the direction of main stress were analyzed. RESULTS: The lumbar spine model was divided into a total of 121 239 nodes, 112 491 units. It could objectively reflect the true anatomy of lumbar spine and its biomechanical behavior and obtain the end-plate images under different stress. The stress distribution on the lumbar intervertebral disc (L(3)-L(4)) under the axial, lateral flexion and extension stress, and the displacement trace of the corresponding processus articularis were analyzed. CONCLUSION: It is helpful to analyze the stress distribution of lumbar spine and units displacement in static stress loading in the clinical research of lumbar spine injury and the distribution of internal stress.

[Strategy and analysis of early management on ninety multiple trauma patients].

OBJECTIVE: To explore the therapeutical effects through investigating the results of multiple traumas treated by means of damage control surgery (DCS) or early total care (ETC). METHODS: The clinical data of 90 patients with severe multiple traumas admitted from June 2001 to June 2007 were analyzed retrospectively. Forty-five patients were classified to the DCS group, 45 patients to the ETC group as a control. These severely injured patients were selected with an ISS > 25 points. RESULTS: Between the two groups, the recovery time of clearance of lactic acid, body temperature, prothrombin time (PT) and activated partial thromboplastin time (APTT) of DCS group was significantly shorter than that of ETC group (P < 0.05). Incidence of complications and mortality in DCS group was significant less than that in ETC group (P < 0.05). No significant differences existed in the volume of bleeding and duration of surgery (P > 0.05). CONCLUSIONS: The concept of DCS could reduce multiple traumas patients' mortality rate and incidence of complications. The indication of DCS should be the combination of physical feature, mechanism and severity of injuries.

Dithiocarbamate-inspired side chain stapling chemistry for peptide drug design.

Two major pharmacological hurdles severely limit the widespread use of small peptides as therapeutics: poor proteolytic stability and membrane permeability. Importantly, low aqueous solubility also impedes the development of peptides for clinical use. Various elaborate side chain stapling chemistries have been developed for α-helical peptides to circumvent this problem, with considerable success in spite of inevitable limitations. Here we report a novel peptide stapling strategy based on the dithiocarbamate chemistry linking the side chains of residues Lys(i) and Cys(i + 4) of unprotected peptides and apply it to a series of dodecameric peptide antagonists of the p53-inhibitory oncogenic proteins MDM2 and MDMX. Crystallographic studies of peptide-MDM2/MDMX complexes structurally validated the chemoselectivity of the dithiocarbamate staple bridging Lys and Cys at (i, i + 4) positions. One dithiocarbamate-stapled PMI derivative, DTCPMI, showed a 50-fold stronger binding to MDM2 and MDMX than its linear counterpart. Importantly, in contrast to PMI and its linear derivatives, the DTCPMI peptide actively traversed the cell membrane and killed HCT116 tumor cells in vitro by activating the tumor suppressor protein p53. Compared with other known stapling techniques, our solution-based DTC stapling chemistry is simple, cost-effective, regio-specific and environmentally friendly, promising an important new tool for the development of peptide therapeutics with improved pharmacological properties including aqueous solubility, proteolytic stability and membrane permeability.

A Highly Efficient Synthesis of Polyubiquitin Chains.

A robust, microwave-assisted, highly efficient, solid-phase peptide synthesis method for preparing isopeptide-linked 62-mer and 76-mer isoubiquitins and polyubiquitin is developed. The strategy avoids the use of costly resins and pseudoprolines, and the isopeptide-linked building blocks can be assembled with high initial purity within 1 day. All seven diubiquitins are successfully synthesized on a multi-milligram scale; a four-segment, three-ligation method is used to obtain a K33-/K11-linked mixed triubiquitin in excellent yield. Circular dichroism and crystallographic analyses are used to verify the structures of the well-folded, synthetic polyubiquitin chains. The facile synthetic strategy is expected to be generally applicable for the rapid synthesis of isopeptide-linked isoUbs and to pave the way for the study of longer polyubiquitin chains.