兔骨髓诱导的内皮细胞和纤维蛋白凝胶体外共培养

目的将兔骨髓诱导的内皮细胞接种于纤维蛋白凝胶内并观察其生长增值情况。方法梯度离心分离兔骨髓单个核细胞（MNCs）,直接向内皮细胞诱导培养,传代培养至第2代细胞,倒置显微镜观察细胞形态及生长特点,免疫组化鉴定;以10^5密度接种于纤维蛋白凝胶中培养并定期观察细胞生长情况,单纯细胞及胶原内的细胞分别用MTT法检测其生长并绘制生长曲线,胶原切片并作苏木精．伊红染色观察细胞在胶原内的生长。结果诱导的内皮细胞为铺路石样。呈单层生长,第2代细胞CD31、八因子相关抗原免疫组化均为阳性,摄取低密度脂蛋白和结合凝集素实验均阳性;细胞在纤维蛋白凝胶内成立体生长,细胞在纤维蛋白凝胶内3d后成梭形铺开,6d后自发形成管腔样结构;细胞在纤维蛋白凝胶内生长曲线成“s”形。结论骨髓诱导的内皮细胞在纤维蛋白凝胶内生长增值良好,纤维蛋白凝胶可以作为接种骨髓诱导的内皮细胞的基质材料。     

Characterization of phenotypes of skeletal Class III malocclusion in Korean adult patients treated with orthognathic surgery using cluster analysis

ObjectivesTo characterize the phenotypes of skeletal Class III malocclusion in adult patients who underwent orthognathic surgery (OGS).Materials and MethodsThe sample consisted of 326 patients with Class III malocclusion treated with OGS (170 men and 156 women; mean age, 22.2 years). Using lateral cephalograms taken at initial visits, 13 angular variables and one ratio cephalometric variable were measured. Using three representative variables obtained from principal components analysis (SNA, SNB, and Björk sum), K-means cluster analysis was performed to classify the phenotypes. Statistical analysis was conducted to characterize the differences in the cephalometric variables among the clusters.ResultsClass III phenotypes were classified into nine clusters from the following four major groups: (1) retrusive maxilla group, clusters 7 and 9 (7.1% and 5.5%; severely retrusive maxilla, normal mandible, severe and moderate hyperdivergent, respectively) and cluster 6 (9.2%; retrusive maxilla, normal mandible, normodivergent); (2) relatively protrusive mandible group, cluster 2 (20.9%; normal maxilla, normal mandible, hyperdivergent); (3) protrusive mandible group, clusters 3 and 1 (11.7% and 15.3%; normal maxilla, protrusive mandible, normodivergent and hyperdivergent, respectively) and clusters 8 and 4 (15.3% and 3.7%; normal maxilla, severe protrusive mandible, normodivergent and hypodivergent, respectively); and (4) protrusive maxilla and protrusive mandible group, cluster 5 (11.4%; protrusive maxilla, severely protrusive mandible, normodivergent). Considerations for presurgical orthodontic treatment and OGS planning were proposed based on the Class III phenotypes.ConclusionsBecause the anteroposterior position of the maxilla and rotation of the mandible by a patient''s vertical pattern determine Class III phenotypes, these variables should be considered in diagnosis and treatment planning for patients who have skeletal Class III malocclusion.     

Evaluation of blood pressure lowering effect by generic and brand-name antihypertensive drugs treatment: a multicenter prospective study in China

Background::Generic drugs are bioequivalent to their brand-name counterparts; however, concerns still exist regarding the effectiveness and safety of generic dr...     

Acute rheumatic fever in adult patients

Acute rheumatic fever (ARF) is considered as a disorder of children, and attacks in adults are usually a recurrence of disease acquired in the child’s life. Although the incidence of ARF in children has a decreasing trend in developed countries, resurgent and sporadic epidemics still occur in adults. The first attacks of ARF in adult patients without a childhood history can lead to a diagnostic dilemma.A medical record review in adults at least 18 years of age with an arthralgia complaint fulfilling 2015 revised Jones criteria was performed from January 1, 2000 to December 31, 2019.Eleven ARF patients were identified, including 8 with initial attacks (6 females aged 26–42 years, 33.9 ± 5.3) and 3 pre-existing valvular heart disease with recurrent attacks (2 females aged 38–52 years, 45.0 ± 7.0). In addition to febrile pharyngitis and migratory polyarthritis in initial attacks, pericarditis was encountered in 1, valvulitis in 2, prolong PR interval in 3 and skin involvement in 2 patients with erythema marginatum and IgA vasculitis. All responded to antibiotics and nonsteroidal anti-inflammatory drugs therapy with normalized clinical and laboratory abnormalities, no new-onset carditis, and no recurrent disease during a long-term follow-up (3.8–19.8 years, 12.7 ± 5.4).A sporadic occurrence of adult ARF is observed in southern Taiwan. This disease should be considered by physicians for the differential diagnosis of febrile pharyngitis with arthritis and/or carditis in adults, even in areas with a low incidence of ARF.     

Imaging Findings of Hepatic Ewing’s Sarcoma on Computed Tomography and Gadobenate Dimeglumine-enhanced Magnetic Resonance Imaging: A Case Report and Literature Review

Ewing’s sarcoma (ES) is a tumor that often occurs in the long bones and rarely arises from visceral organs primarily. Here, we report a case of primary hepatic ES, discuss its computed tomography (CT) and gadobenate dimeglumine-enhanced magnetic resonance (MRI) features. This is the first Chinese and fifth primary hepatic ES case reported, based on a literature review. Imaging examinations showed that the tumor was solid, with necrosis and hemorrhage. Contrast-enhanced images showed that the tumor was hypervascular and especially had heterogeneous signal intensity on hepatobiliary phase MRI images. Intratumoral vessels and vascular invasion were also present.     

Structural conservation among variants of the SARS-CoV-2 spike postfusion bundle

Variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) challenge currently available COVID-19 vaccines and monoclonal antibody therapies due to structural and dynamic changes of the viral spike glycoprotein (S). The heptad repeat 1 (HR1) and heptad repeat 2 (HR2) domains of S drive virus–host membrane fusion by assembly into a six-helix bundle, resulting in delivery of viral RNA into the host cell. We surveyed mutations of currently reported SARS-CoV-2 variants and selected eight mutations, including Q954H, N969K, and L981F from the Omicron variant, in the postfusion HR1HR2 bundle for functional and structural studies. We designed a molecular scaffold to determine cryogenic electron microscopy (cryo-EM) structures of HR1HR2 at 2.2–3.8 Å resolution by linking the trimeric N termini of four HR1 fragments to four trimeric C termini of the Dps4 dodecamer from Nostoc punctiforme. This molecular scaffold enables efficient sample preparation and structure determination of the HR1HR2 bundle and its mutants by single-particle cryo-EM. Our structure of the wild-type HR1HR2 bundle resolves uncertainties in previously determined structures. The mutant structures reveal side-chain positions of the mutations and their primarily local effects on the interactions between HR1 and HR2. These mutations do not alter the global architecture of the postfusion HR1HR2 bundle, suggesting that the interfaces between HR1 and HR2 are good targets for developing antiviral inhibitors that should be efficacious against all known variants of SARS-CoV-2 to date. We also note that this work paves the way for similar studies in more distantly related viruses.

Three previously unknown beta-coronaviruses have emerged in the first two decades of this century: severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003, Middle East respiratory syndrome coronavirus (MERS-CoV) in 2012, and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in late 2019 (1). The most recent outbreak of SARS-CoV-2 that causes coronavirus disease 2019 (COVID-19) has claimed about 6 million lives in 2 y, and several variants of concern have emerged around the globe despite the relatively low mutation rate of coronaviruses (2). Some of these variants pose a challenge to currently available vaccines (3–6), likely due to structural changes of the target of these vaccines (7–11). Hence, there is an urgent need for new antiviral therapeutics (12) that target regions of viruses with conserved structural features that are less likely to be affected by mutations.SARS-CoV, MERS-CoV, and SARS-CoV-2 are enveloped viruses that rely on membrane fusion to deliver RNA to the host cell (13). In each case, the process of viral membrane fusion (14, 15) is mediated by the trimeric viral spike glycoprotein (S) that is cleaved into S1 and S2 subunits by multiple host proteases upon infection (16) (Fig. 1A). S1 recognizes the human angiotensin-converting enzyme 2 (ACE2) receptor and dissociates from S2. Subsequently, S2 undergoes substantial conformational changes that drive membrane remodeling. Similar to other enveloped viruses (14, 15), this process likely proceeds via an intermediate extended state that pulls together the two membranes via the transmembrane domain and fusion peptide of the S2 subunit (17). Two heptad repeat regions, HR1 and HR2, distant from each other in the prefusion S, drive membrane fusion by assembly into a six-helix bundle (18). This HR1HR2 bundle formation is thought to provide the energy for membrane fusion and is therefore a target for therapeutics, as exemplified by peptide inhibitors that disrupt infection by the HIV-1 (19, 20), SARS-CoV (21), MERS-CoV (22), SARS-CoV-2 (23–25), human parainfluenza virus 3 (26), and respiratory syncytial virus (26).Open in a separate windowFig. 1.Mutations of interest in the HR1HR2 bundle of SARS-CoV-2 variants. (A) Schematic diagram of the domain structures of the SARS-CoV-2 spike protein. The N and C termini are labeled on the left and right, respectively. FP, fusion peptide; HR1, heptad repeat 1; HR2, heptad repeat 2; TM, transmembrane region. (B) Locations of the five selected point mutations of SARS-CoV-2 variants (black spheres) and the three mutations of the SARS-CoV-2 Omicron variant (purple spheres) indicated in the crystal structure of the HR1HR2 bundle (PDB ID code 6lxt). Two HR2 residues, R1185 and N1187, that may be affected by the selected mutations are shown as red spheres. The HR1 and HR2 fragments are colored as light blue and light red, respectively. (C) Effects on fusion activity of these mutations. The fusion activity is shown as a percentage (Left)/fold change (Right) relative to that of the wild type (Materials and Methods). The Omicron construct used here for the fusion assay has three mutations—Q954H, N969K, and L981F—in the HR1 portion of the HR1HR2 bundle, but not other mutations from different regions of the spike found in the Omicron variant. *P < 0.05, **P < 0.01, ***P < 0.001, by a Student’s t test.Despite the established value of inhibitors targeting formation of the HR1HR2 bundle, the structural plasticity of this bundle upon mutation is largely unknown. Comparison with distantly related viruses suggests that the overall architecture is maintained despite vast differences in primary sequence (SI Appendix, Fig. S1). To what degree does the structure of the HR1HR2 bundle change upon mutation? To address this question, we surveyed mutations of all currently known variants (including Omicron) of SARS-CoV-2 S in the postfusion HR1HR2 bundle, selected eight mutations of potential interest, and investigated their effects on structure and function.Structural characterization of the HR1HR2 bundle has proven surprisingly challenging. To date, two successful approaches for determining structures of the HR1HR2 bundle have been employed. First, several HR1HR2 structures with the HR1 and HR2 domains synthetically linked were determined by X-ray crystallography (2.9 Å, Protein Data Bank [PDB] ID code 6lxt; 1.5 Å, PDB ID code 6m1v) (23, 25). Second, a sample of postfusion S2 was generated from a recombinant source (mammalian HEK-293F cells) expressing full-length S; as such, multiple states of S undergoing spontaneous transition from the prefusion to the postfusion state were present in the sample and the postfusion structure was determined by single-particle cryogenic electron microscopy (cryo-EM) (3.0 Å, PDB ID code 6xra) (18). Although the structures of the postfusion HR1HR2 bundle are similar, there are differences between these structures and the local resolution is quite variable or limited. More importantly, neither approach is particularly suited for efficient structure determination of multiple mutants at high resolution. Therefore, we decided to develop a platform for using single-particle cryo-EM to efficiently determine structures of HR1HR2 bundles at atomic resolution.The postfusion HR1HR2 bundle of SARS-CoV-2 is a 115 × 25 × 25 Å bundle consisting of six helices (PDB ID code 6lxt) (23). Its molecular weight is 40 kDa, close to the theoretical minimum size needed to achieve a reconstruction with near-atomic resolution by cryo-EM (27). To our knowledge, it has not yet been possible to determine structures of individual proteins <50 kDa to high resolution, with exception in the case of multimers (28, 29) or small RNA molecules (30). In addition, efforts extending the resolution limit of cryo-EM have largely focused on globular proteins (28, 29, 31, 32), perhaps because fibrous samples are more flexible, more susceptible to the issue of preferred orientation, and require thicker ice to bury the entire particle—all of which inevitably increase noise in the already extremely low-contrast and hard-to-align images. To overcome the size limit of single-particle cryo-EM, two strategies have been employed to increase the effective mass of the target protein, e.g. using antibodies/nanobodies/legobodies (33, 34) and molecular scaffolds (35–38). Since developing antibodies/nanobodies/legobodies can be time-consuming we resorted to the molecular scaffold approach. We first attempted to use existing scaffolds but were unable to engineer a linkage ensuring proper HR1HR2 bundle formation. We therefore designed a scaffold to efficiently determine structures of the postfusion HR1HR2 bundle and its mutants to near-atomic resolution by single-particle cryo-EM.Our high-resolution wild-type structure of the HR1HR2 bundle resolves uncertainties in some side-chain positions present in prior structures. Our HR1HR2 structures of SARS-CoV-2 variants reveal an overall architecture that is highly conserved, with only side-chain rearrangement for five point mutations and, for the Omicron variant containing three mutations in HR1, a slight shift of the HR2 backbone in a nonhelical region that interacts with HR1. These results suggest that interactions between HR1 and HR2 are excellent targets for disruption by broadly efficacious antiviral inhibitors. Moreover, our approach can be directly used to study the binding of potential HR2-based peptide inhibitors and adapted to study the postfusion bundles of other coronaviruses or other structurally similar viruses.     

Self-adaptive integration of photothermal and radiative cooling for continuous energy harvesting from the sun and outer space

The sun (∼6,000 K) and outer space (∼3 K) are two significant renewable thermodynamic resources for human beings on Earth. The solar thermal conversion by photothermal (PT) and harvesting the coldness of outer space by radiative cooling (RC) have already attracted tremendous interest. However, most of the PT and RC approaches are static and monofunctional, which can only provide heating or cooling respectively under sunlight or darkness. Herein, a spectrally self-adaptive absorber/emitter (SSA/E) with strong solar absorption and switchable emissivity within the atmospheric window (i.e., 8 to 13 μm) was developed for the dynamic combination of PT and RC, corresponding to continuously efficient energy harvesting from the sun and rejecting energy to the universe. The as-fabricated SSA/E not only can be heated to ∼170 °C above ambient temperature under sunshine but also be cooled to 20 °C below ambient temperature, and thermal modeling captures the high energy harvesting efficiency of the SSA/E, enabling new technological capabilities.

Heating and cooling are two kinds of significant end uses of thermal energy in society, which exist in various conditions (e.g., space/water heating, space cooling, and industrial processes) and account for 51% of the total final energy consumption (1). For example, the heating and cooling of buildings are responsible for nearly 48% of the building energy consumption, increasing to be the largest individual energy expense (2). Therefore, heat and cool harvesting relying on clean techniques from renewable energy resources has drawn remarkable attention from fields of engineering to material science because it has considerable potential for global energy conservation and greenhouse emission reduction. Thermodynamically, any heat transportation and work-generation process requires a temperature gradient. The hot sun (∼6,000 K) and cold outer space (∼3 K) are the ultimate heat source and heat sink for the Earth. Theoretical analysis reveals that maximal output work can be extracted from nonreciprocal systems based on the temperature difference between the sun and Earth (∼300 K) with an ultimate solar energy harvesting efficiency limit of 93.3%, while a maximal work of 153.1 W·m⁻² can also be obtained on the basis of temperature difference between the Earth and outer space (3, 4). Thus, the sun and outer space are two significant renewable thermodynamic resources for the Earth, which can be effectively utilized for clean heat and cool collection.Photothermal (PT) is a widely used solar thermal collection method that employs solar absorbers to capture solar photons and convert them to heat. Thermal analysis reveals that a good candidate for a solar absorber should have high solar absorptivity and low thermal emissivity simultaneously for efficient solar thermal collection. Various materials, including multilayer metal/ceramic films (5, 6), photonic crystals (7, 8), and metamaterials (9, 10), have been developed for spectrally selective solar absorbers and have been used for real-world applications. Meanwhile, radiative cooling (RC) has re-elicited considerable interest in recent years because it can passively provide clean cooling without any extra energy input (11–14). The waste heat of terrestrial objects can be continuously pumped into the cold outer space, relying on the transparent atmospheric window (i.e., 8 to 13 μm). So, high emissivity within the atmospheric window of materials is necessary for efficient RC, and excellent solar reflection is also important for RC under sunshine. Thus, different materials with the tailored spectrum, such as photonic structures (15–17), structure materials (18), energy-saving paints (19–21), and even metamaterials (22–24), have been reported for passive cooling. On the potential application level, RC implementations also span a range of fields, including passive cooling of buildings (25–27), thermal management of textiles and color surfaces (28–30), atmospheric water harvesting (31), and thermoelectric generation (32, 33). Although the reported PT and RC can generate heat and cold with high efficiency through different spectrally selective materials, most of the approaches are static and monofunctional, which can only provide heating or cooling under sunlight or darkness. Therefore, the dynamical integration of PT and RC for continuously efficient heat and cool harvesting is a new topic for the energy exploitation of the sun and outer space. The tunable combination of PT and RC hybrid utilization has been recently proposed, but mechanical methods such as switching (e.g., flip action) a PT absorber and an RC emitter manually (34) or changing the optical properties of the materials through extra force stimuli (35) are preferred.Herein, a smart strategy for the dynamic combination of daytime PT and nighttime RC is proposed, corresponding to continuously efficient energy harvesting from the sun and rejecting energy to the universe. A spectrally self-adaptive absorber/emitter (SSA/E) with solar absorption of over 0.8 and emissivity modulation capability of regulating from broadband emissivity of 0.25 within the mid-infrared (MIR) region to the selective high emissivity of 0.75 within the atmospheric window is designed and fabricated for the proof of the concept. Outdoor thermal experimental results demonstrate that the SSA/E can be heated to ∼170 °C above ambient temperature in the daytime PT mode and passively cooled to ∼20 °C below ambient temperature in the nighttime RC mode. Moreover, the heat and cool energy gains of the SSA/E system are respectively predicted to be 78% and 103% larger than those of the reference system that combines static and monofunctional PT absorber and RC emitter.