血小板浓缩生长因子对正畸大鼠牙移动及牙周组织CCR-1和HO-1表达的影响

摘要 目的：研究血小板浓缩生长因子（concentrated growth factors, CGF）对正畸大鼠牙移动及牙周组织CC类趋化因子受体-1（CC chemokine receptor-1, CCR-1）及血红素氧合酶-1（heme oxygenase-1, HO-1）表达的影响。方法：150只大鼠随机分为对照组和观察组,每组75只,两组均建立正畸牙移动模型,观察组每日腹腔注射10 % CGF生理盐水溶液,对照组注射等体积生理盐水溶液,共注射14 d。分别检测第0 d、1 d、3 d、7 d和14 d两组大鼠的牙移动距离,TRAP染色并进行破骨细胞计数,检测牙周组织中CCR1和HO-1 mRNA的相对表达。结果：随正畸力作用时间的延长,两组大鼠牙移动距离均显著增长、破骨细胞计数显著增多、牙周组织中CCR1和HO-1 mRNA的相对表达均显著升高（P＜0.05）;对照组与观察组在第1 d和第3 d时牙移动距离相比无统计学差异（P＞0.05）,而在第7 d开始,观察组牙移动距离显著长于对照组（P＜0.05）;观察组大鼠破骨细胞计数和牙周组织中CCR1和HO-1 mRNA的相对表达均显著高于同时间点的对照组（P＜0.05）。结论：CGF可促进正畸大鼠牙移动,增加破骨细胞数量,从而促进正畸牙移动和牙周组织重建,其机制可能与增加牙周组织中CCR1和HO-1的表达水平有关。     

Crystal structure and characterization of coiled-coil domain of the transient receptor potential channel PKD2L1

The cation-permeable channel PKD2L1 forms a homomeric assembly as well as heteromeric associations with both PKD1 and PKD1L3, with the cytoplasmic regulatory domain (CRD) of PKD2L1 often playing a role in assembly and/or function. Our previous work indicated that the isolated PKD2L1 CRD assembles as a trimer in a manner dependent on the presence of a proposed oligomerization domain. Herein we describe the 2.7 Å crystal structure of a segment containing the PKD2L1 oligomerization domain which indicates that trimerization is driven by the β-branched residues at the first and fourth positions of a heptad repeat (commonly referred to as “a” and “d”) and by a conserved R-h-x-x-h-E salt bridge motif that is largely unique to parallel trimeric coiled coils. Further analysis of the PKD2L1 CRD indicates that trimeric association is sufficiently strong that no other species are present in solution in an analytical ultracentrifugation experiment at the lowest measurable concentration of 750 nM. Conversely, mutation of the “a” and “d” residues leads to formation of an exclusively monomeric species, independent of concentration. Although both monomeric and WT CRDs are stable in solution and bind calcium with 0.9 μM affinity, circular dichroism studies reveal that the monomer loses 25% more α-helical content than WT when stripped of this ligand, suggesting that the CRD structure is stabilized by trimerization in the ligand-free state. This stability could play a role in the function of the full-length complex, indicating that trimerization may be important for both homo- and possibly heteromeric assemblies of PKD2L1.     

Coupling σ Factor Conformation to RNA Polymerase Reorganisation for DNA Melting

ATP-driven remodelling of initial RNA polymerase (RNAP) promoter complexes occurs as a major post recruitment strategy used to control gene expression. Using a model-enhancer-dependent bacterial system (σ⁵⁴-RNAP, Eσ⁵⁴) and a slowly hydrolysed ATP analogue (ATPγS), we provide evidence for a nucleotide-dependent temporal pathway leading to DNA melting involving a small set of σ⁵⁴-DNA conformational states. We demonstrate that the ATP hydrolysis-dependent remodelling of Eσ⁵⁴ occurs in at least two distinct temporal steps. The first detected remodelling phase results in changes in the interactions between the promoter specificity σ⁵⁴ factor and the promoter DNA. The second detected remodelling phase causes changes in the relationship between the promoter DNA and the core RNAP catalytic β/β′ subunits, correlating with the loading of template DNA into the catalytic cleft of RNAP. It would appear that, for Eσ⁵⁴ promoters, loading of template DNA within the catalytic cleft of RNAP is dependent on fast ATP hydrolysis steps that trigger changes in the β′ jaw domain, thereby allowing acquisition of the open complex status.     

A Single-Cell Atlas of Tumor-Infiltrating Immune Cells in Pancreatic Ductal Adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies with limited treatment options. To guide the design of more effective immunotherapy strategies, mass cytometry was employed to characterize the cellular composition of the PDAC-infiltrating immune cells. The expression of 33 protein markers was examined at the single-cell level in more than two million immune cells from four types of clinical samples, including PDAC tumors, normal pancreatic tissues, chronic pancreatitis tissues, and peripheral blood. Based on the analyses, we identified 23 distinct T-cell phenotypes, with some cell clusters exhibiting aberrant frequencies in the tumors. Programmed cell death protein 1 (PD-1) was extensively expressed in CD4⁺ and CD8⁺ T cells and coexpressed with both stimulatory and inhibitory immune markers. In addition, we observed elevated levels of functional markers, such as CD137L and CD69, in PDAC-infiltrating immune cells. Moreover, the combination of PD-1 and CD8 was used to stratify PDAC tumors from The Cancer Genome Atlas database into three immune subtypes, with S1 (PD-1⁺CD8⁺) exhibiting the best prognosis. Further analysis suggested distinct molecular mechanisms for immune exclusion in different subtypes. Taken together, the single-cell protein expression data depicted a detailed cell atlas of the PDAC-infiltrating immune cells and revealed clinically relevant information regarding useful cell phenotypes and targets for immunotherapy development.     

Prognostic value of the expression of chemokines and their receptors in regional lymph nodes of melanoma patients

Chemokines and their receptors have been reported to drive immune cells into tumours or to be directly involved in the promotion or inhibition of the development of tumours. However, their expression in regional lymph node (LN) tissues in melanoma patients remains unknown. The present study investigated the relationship between the expression of mRNA of chemokines and their receptors and clinicopathology of the regional LN tissues of skin cutaneous melanoma (SKCM) patients available in The Cancer Genome Atlas. The relationship between chemokines and their receptors and the composition of immune cells within the tumour was analysed. In SKCM regional LN tissues, the high expression of 32 types of chemokines and receptors, namely CCL2, 4-5, 7-8, 13, 22-25, CCR1-9, CXCL9-13, 16, CXCR3, 5, 6, XCL1-2 and XCR1 in LN was associated with favourable patient prognosis. Conversely, high expression of CXCL17 was an indicator of poor prognosis. The expression of mRNA for CXCL9-11, 13, CXCR3, 6, CCL2, 4, 5, 7, 8, 25, CCR1, 2, 5, and XCL1, 2 in regional LN tissues was positively correlated with the fraction of CD8-positive T cells and M1 macrophages, and was negatively correlated with M0 macrophages. CCR4, 6-9, CCL13, 22, 23 and XCR1 were positively correlated with the fraction of memory B cells and naive T cells, and negatively correlated with M0 macrophages and resting mast cells, suggesting that chemokines and their receptors may affect the prognosis of patients by guiding immune cells into the tumour microenvironment to eliminate tumour cells.     

Polymorphisms of transforming growth factor beta 1 (RS#1800468 and RS#1800471) and esophageal squamous cell carcinoma among Zhuangese population,China

Epidemiological evidence has shown two polymorphisms (namely RS#1800468G > A and RS#1800471G > C) of transforming growth factor-beta 1 (TGF-β1) gene may be involved in the cancer development. However, their role in the carcinogenic process of esophageal squamous cell carcinoma (ESCC) has been less well elaborated. We conducted a hospital-based case-control study including 391 ESCC cases and 508 controls without any evidence of tumors to evaluate the association between these two polymorphisms and ESCC risk and prognosis for Zhuangese population by means of polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and amplification refractory mutation system (ARMS)-PCR techniques. We found that individuals with the genotypes with RS#1800471 C allele (namely RS#1800471-GC or -CC) had an increased risk of ESCC than those without above genotypes (namely RS#1800471-GG, adjusted odds ratio 3.26 and 5.65, respectively). Further stratification analysis showed that this polymorphism was correlated with tumor histological grades and TNM (tumor, node, and metastasis) stage, and modified the serum levels of TGF-β1. Additionally, RS#1800471 polymorphism affected ESCC prognosis (hazard ratio, 3.40), especially under high serum levels of TGF-β1 conditions. However, RS#1800468 polymorphism was not significantly related to ESCC risk. These findings indicated that TGF-β1 RS#1800471G > C polymorphism may be a genetic modifier for developing ESCC in Zhuangese population.     

The Conformation and Orientation of a 27-Residue CCR5 Peptide in a Ternary Complex with HIV-1 gp120 and a CD4-Mimic Peptide

Interaction of CC chemokine receptor 5 (CCR5) with the human immunodeficiency virus type 1 (HIV-1) gp120/CD4 complex involves its amino-terminal domain (Nt-CCR5) and requires sulfation of two to four tyrosine residues in Nt-CCR5. The conformation of a 27-residue Nt-CCR5 peptide, sulfated at Y10 and Y14, was studied both in its free form and in a ternary complex with deglycosylated gp120 and a CD4-mimic peptide. NMR experiments revealed a helical conformation at the center of Nt-CCR5(1-27), which is induced upon gp120 binding, as well as a helical propensity for the free peptide. A well-defined structure for the bound peptide was determined for residues 7-23, increasing by 2-fold the length of Nt-CCR5's known structure. Two-dimensional saturation transfer experiments and measurement of relaxation times highlighted Nt-CCR5 residues Y3, V5, P8-T16, E18, I23 and possibly D2 as the main binding determinant. A calculated docking model for Nt-CCR5(1-27) suggests that residues 2-22 of Nt-CCR5 interact with the bases of V3 and C4, while the C-terminal segment of Nt-CCR5(1-27) points toward the target cell membrane, reflecting an Nt-CCR5 orientation that differs by 180° from that of a previous model. A gp120 site that could accommodate ^CCR5Y3 in a sulfated form has been identified. The present model attributes a structural basis for binding interactions to all gp120 residues previously implicated in Nt-CCR5 binding. Moreover, the strong interaction of sulfated ^CCR5Tyr14 with ^gp120Arg440 revealed by the model and the previously found correlation between E322 and R440 mutations shed light on the role of these residues in HIV-1 phenotype conversion, furthering our understanding of CCR5 recognition by HIV-1.