Comprehensive analysis of RHD splicing transcripts reveals the molecular basis for the weak anti-D reactivity of Del -red blood cells.

BACKGROUND: Of the Rh blood type, Del is a rare variant that elicits the weakest anti-D reactivity. In this study, we revisit the genetic changes of Del allele and characterise the RHD splicing transcripts to realise the molecular basis of Del formation in the Taiwanese population. STUDY DESIGN AND METHODS: The RHD exons from Del and D-positive individuals were amplified by polymerase chain reaction (PCR) using different primer pairs followed by sequencing analyses. In addition, full-length RHD transcripts were reversed transcribed and amplified by nested-PCR. The type and frequency of the RHD splicing transcripts were analysed after sequencing the PCR products that were subcloned into a cloning vector. RESULTS: All Del individuals had a characteristic 1227G>A mutation. No deletion of the exon sequences was found. At least nine types of RHD splicing transcripts including exons 7/8/9 deletion, 7/9 deletion, 8/9 deletion, 9 deletion, 2/3/7/9 deletion, 2/3/7/8/9 deletion, exons 7/8/9 deletion with replacement of exon 3 with RHCE exon 3, exon 9 deletion with cryptic insertion of 170 bp of intron 7 and exons 7/8/9 deletion with cryptic insertion of 117 bp of intron 3 were identified in the Del -RBC. These aberrant splicing transcripts led to production of frame shift or truncated D antigen. Notably, no full-length RHD transcript was identified in the Del -RBC. CONCLUSION: The RHD 1227G>A mutation contributes to the molecular basis of Del phenotype in the Taiwanese population. The point mutation results in aberrant frame shift or exon deletion transcripts and generates D protein with weak antigen presenting function.

Genetic and mechanistic evaluation for the weak A phenotype in Ael blood type with IVS6 + 5G>A ABO gene mutation.

BACKGROUND AND OBJECTIVES: Ael is a rare blood type that is characterized by weak agglutination of RBCs when reacts with anti-A antibody in adsorption-elution test. Although IVS6 + 5G→A mutation is known to associate with the Ael blood type, genetic and mechanistic evaluation for the weak agglutination of Ael with IVS6 + 5G→A mutation has not yet been completely addressed. MATERIALS AND METHODS: In this study, five cases of confirmed Ael individuals were analysed. The cDNAs for the A(el) alleles were obtained by cloning method for sequence analyses. The erythroleukemia K562 cells were used as the cell study model and were transfected with the A(el) expression construct. Flow cytometry analysis was then performed to determine the levels of surface antigen expression. RESULTS: The results indicated that IVS6 + 5G→A attributes to all cases of Ael . RT-PCR analyses revealed the presence of at least 10 types of aberrant A(el) splicing transcripts. Most of the transcripts caused early termination and produced non-functional protein during translation. Nevertheless, the transcript without exons 5-6 was predicted to generate functional Ael glycosyltransferase lacking 57 amino acids at the N-terminal segment. When the exons 5-6 deletion transcript was stably expressed in the K562 cells, weak agglutination of the cells can be induced by adding anti-A antibody followed by adsorption-elution test. CONCLUSION: This study demonstrates that aberrant splicing of A transcripts contributes to weak A expression and the weak agglutination of Ael -RBCs, adding to the complexity for the regulatory mechanisms of ABO gene expression.

HLA-A*31:01 and different types of carbamazepine-induced severe cutaneous adverse reactions: an international study and meta-analysis.

HLA-A*31:01 was reported to be associated with carbamazepine (CBZ)-induced severe cutaneous adverse reactions (SCAR), including drug reaction with eosinophilia and systemic symptoms (DRESS), Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN). We conducted an international study using consensus diagnosis criteria to enroll a total of 93 patients with CBZ-SCAR from Europe or Asia. We found that HLA-A*31:01 showed a significant association with CBZ-DRESS in Europeans (P<0.001; odds ratio (OR) (95% confidence interval (CI))=57.6 (11.0-340)), and the strong association was also found in Chinese (P<0.001; OR (95% CI)=23.0 (4.2-125)). However, HLA-A*31:01 had no association with CBZ-SJS/TEN in neither Chinese nor Europeans. By comparison, HLA-B*15:02 showed a strong association with CBZ-SJS/TEN in Chinese (P<0.001, OR (95% CI)=58.1 (17.6-192)). A meta-analysis of this and other published studies confirmed that in all populations, HLA-A*31:01 had an extremely strong association with CBZ-DRESS (P<0.001, a pooled OR (95% CI)=13.2 (8.4-20.8)), but a much weaker association with CBZ-SJS/TEN (P=0.01, OR (95% CI)=3.94 (1.4-11.5)). Our data revealed that HLA-A*31:01 is a specific predictor for CBZ-DRESS but not for CBZ-SJS/TEN. More studies are needed to investigate the genetic determinant of CBZ-SJS/TEN in Europeans. Considering the potential clinical utility, the cost-effectiveness of the combined HLA-A*31:01 and HLA-B*15:02 genetic test to prevent CBZ-SCAR in Chinese needs further investigation.

[Effect of digital intraoral full-arch scan strategies on scan time and accuracy on conditions of intraoral head-simulator].

Objective: To comparatively evaluate the accuracy and the scan time of three full-arch scan strategies on the head-simulator, to explore a full-arch scan strategy with better clinical operability and high accuracy. Methods: A cross-controlled study design was used. A model with melamine-formaldehyde resin teeth and silica gel gingiva of an upper dental arch which can be fixed on a head simulator was scanned with an optical scanner (ATOS Core) in order to obtain the standard tessellation language (STL) dataset as reference. Intraoral scans were performed on the model fixed on the head simulator with four intraoral scanners (IOS) [A (TRIOS 3), B (CS 3600), C (CEREC Omnicam), D (iTero)]. The STL datasets were obtained from each of the four different IOS systems by using three scan strategies (scan strategies 1, 2 and 3 were composed of 10, 5 and 7 paths respectively) all by one attending doctor with 3 years of intraoral scanning experience. For each scanner and each scan strategy, nine scans were acquired. And the scan time was recorded for each scan. Following the scan strategy, the scan path was completed to obtain a full-arch digital model, and the scan time was recorded as full-arch scan time. Complementary scans were performed to fill the missing image, and this scan time was recorded as complementary scan time. The total scan time was obtained by adding full-arch scan time and complementary scan time. Through the Geomagic Wrap software, the three-dimensional (3D) models were overlaid by best fit alignment function and compared to obtain the root mean square values of the discrepancies by 3D compare function. The intraoral scanning datasets were compared with the reference for trueness. The nine intraoral scanning datasets were cross compared with same scan strategy and same intraoral scanner for precision. Results: There were no significant differences among the three scan strategies for trueness (P>0.05), while the differences among the three scan strategies for precision were affected by difference IOSs (P<0.05), and only scan strategy 3 showed the highest precision with all the four IOS. The full-arch scan time of scan strategies 1, 2 and 3 were (130±24), (72±17) and (90±19) s respectively (P<0.05). For complementary scan time, scan strategy 2 [(50±24) s] took longer time than scan strategy 1 [(26±18) s] and scan strategy [(25±21) s] (P<0.05), while no significant differences between the latter two (P>0.05). For total scan time, scan strategy 1 [(156±31) s] took longer time than scan strategy 2 [(122±30) s ] and scan strategy 3 [(115±29) s ] (P<0.05), while no significant differences between the latter two (P>0.05). Conclusions: Full-arch scanning on the head-simulator with scan strategy 3 which can obtain scanning datasets with high accuracy, was more convenient to operate and took shorter scan time, and is generally suitable for intraoral scanners commonly used in clinic.

Elevated production of B cell chemokine CXCL13 is correlated with systemic lupus erythematosus disease activity.

INTRODUCTION: B lymphocyte chemoattractant (BLC/CXCL13), a CXC chemokine, is involved in B1 and B2 cell trafficking for the activation of autoreactive T helper (Th) cells and autoantibody production in target organs during the development of lupus. CXCL13 can induce the trafficking of CXCR5+ T lymphocyte subset designated as follicular helper T lymphocytes (T(FH)) which are specifically involved in autoantibody production. MATERIALS AND METHODS: We herein measured the plasma concentrations of CXCL13, B-cell-activating factor of the TNF family (BAFF), and T(FH)-cells-related cytokine IL-21 and cell surface expression of T(FH)-related receptor CXCR5 and IL-21R on CD4+Th and CD19+B cells in 35 systemic lupus erythematosus (SLE) patients and 23 sex- and age-matched control subjects (NC) using enzyme-linked immunosorbent assay and flow cytometry, respectively. RESULTS AND DISCUSSION: Plasma CXCL13, BAFF, and IL-21 concentrations were significantly higher in SLE patients than NC group (all p < 0.0001). Increase in CXCL13 concentration correlated positively and significantly with SLEDAI score in SLE patients (r = 0.399, p = 0.032). Cell surface expression of CXCR5 on Th and B cells and IL-21R on B cells was however significantly lower in SLE patients than controls (both p < 0.01). It may indicate that most differentiated T(FH) cells migrate out from circulation into lymphoid organ upon activation during the disease development of SLE. CONCLUSIONS: The above results suggest that the elevated production of CXCL13, BAFF, and IL-21 may be associated with the function of T(FH) for the immunopathogenesis in SLE, and CXCL13 may serve as a potential disease marker of SLE.

Activation profile of Toll-like receptors of peripheral blood lymphocytes in patients with systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease associated with aberrant activation of T and B lymphocytes for the production of inflammatory cytokines and autoreactive antibodies. Animal studies of SLE have indicated that Toll-like receptors (TLR) are important in the pathogenesis of murine lupus. In the present clinical study, differential protein expressions of TLR-1-9 of monocytes and different lymphocyte subsets from patients with SLE and normal control subjects were determined by flow cytometry. Results showed that the expression of intracellular TLRs (TLR-3, -8, -9) and extracellular TLRs (TLR-1, -2, -4, -5, -6) were elevated in monocytes, CD4(+) T lymphocytes, CD8(+) T lymphocytes and B lymphocytes of SLE patients compared to control subjects (all P < 0.001). Moreover, cell surface expression of TLR-4 on CD4(+) T lymphocytes and CD8(+) T lymphocytes, and TLR-6 on B lymphocytes, were correlated positively with SLE disease activity index (SLEDAI) (TLR-4 on CD4(+) T lymphocytes and CD8(+) T lymphocytes: r = 0.536, P = 0.04; r = 0.713, P = 0.003; TLR-6 in B lymphocytes: r = 0.572, P = 0.026). In concordance with the above results, there is an observable increased relative induction (%) of inflammatory cytokine interleukin (IL)-1beta, IL-6, IL-10 and IL-12, chemokines CCL2, CXCL8, CCL5 and CXCL10 from peripheral blood mononuclear cells (PBMC) upon differential stimulation by PolyIC (TLR-3 ligand), lipopolysaccharide (TLR-4 ligand), peptidoglycan (TLR-2 ligand), flagellin (TLR-5 ligand), R837 (TLR-7 ligand) and CpG DNA (TLR-9 ligand) in SLE patients compared to controls. These results suggest that the innate immune response for extracellular pathogens and self-originated DNA plays immunopathological roles via TLR activation in SLE.

Two-dimensional geometry of spin excitations in the high-transition-temperature superconductor YBa2Cu3O6+x.

The fundamental building block of the copper oxide superconductors is a Cu4O4 square plaquette. The plaquettes in most of these materials are slightly distorted to form a rectangular lattice, for which an influential theory predicts that high-transition-temperature (high-T(c)) superconductivity is nucleated in 'stripes' aligned along one of the axes. This theory received strong support from experiments that indicated a one-dimensional character for the magnetic excitations in the high-T(c) material YBa2Cu3O6.6 (ref. 4). Here we report neutron scattering data on 'untwinned' YBa2Cu3O6+x crystals, in which the orientation of the rectangular lattice is maintained throughout the entire volume. Contrary to the earlier claim, we demonstrate that the geometry of the magnetic fluctuations is two-dimensional. Rigid stripe arrays therefore appear to be ruled out over a wide range of doping levels in YBa2Cu3O6+x, but the data may be consistent with liquid-crystalline stripe order. The debate about stripes has therefore been reopened.

[Evaluation of Schneiderian membrane state using fiber optic endoscope during maxillary sinus floor elevation with lateral window].

Objective: To observe the status of the sinus membrane using fiber optic endoscope during the lateral window approach sinus floor elevation to provide a reference for clinicians when evelvating the sinus mucoperiosteum. Methods: Sixty-six patients (72 sides) who underwent maxillary sinus floor elevation in Beijing Ruicheng Stomatology Hospital from September 2016 to December 2019 were selected, including 40 males and 26 females, aged 26-80 years old [(56.2±11.5) years]. And fiber optic endoscopy was used to observe the maxillary mucoperiosteum during the operation. Results: The status of maxillary sinus mucoperiosteal during lateral window approach sinus floor elevation can be divided into four categories: ① Class Ⅰ, complete periosteal, no damage to mucoperiosteum; ②Class Ⅱ, periosteal injury, unexposed laminae propria; ③Class Ⅲ, periosteal Rupture, exposed lamina propria; ④ Class Ⅳ, mucoperiosteum perforation, rupture of periosteum, lamina propria and epithelial layer. A total of 72 operations were performed, including 18 cases of class I, 28 cases of class Ⅱ, 4 cases of class Ⅲ, and 22 cases of class Ⅳ. Conclusions: The status of maxillary sinus mucoperiosteal during lateral window approach sinus floor elevation can be divided into four categories. Fiberoptic endoscopy as a clinical auxiliary examination method can improve the operator's control of the status of the maxillary sinus membrane and assist the peeling of the mucosa.

Activation profile of intracellular mitogen-activated protein kinases in peripheral lymphocytes of patients with systemic lupus erythematosus.

INTRODUCTION: Systemic lupus erythematosus (SLE) is a systemic autoimmune disease associated with aberrant activation of T and B lymphocytes. Abnormal activation of intracellular signaling molecules in lymphocytes by inflammatory cytokines can instigate the inflammation in SLE. MATERIALS AND METHODS: The activation of extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) in inflammatory cytokine IL-18-activated monocytes, CD4+ T helper (Th) lymphocytes, CD8+ T lymphocytes, and CD19+ B lymphocytes in 22 SLE patients and 20 sex- and age-matched control subjects were measured by flow cytometry. RESULTS AND DISCUSSION: The basal expressions of phospho-p38 MAPK in CD4+ T lymphocytes, CD8+ T lymphocytes, and B lymphocytes were significantly higher in SLE patients than controls (all p<0.05). The expression of phospho-p38 MAPK in CD4+ T lymphocytes, CD8+ T lymphocytes and B lymphocytes, and phospho-JNK in CD8+ T lymphocytes and B lymphocytes was also significantly elevated in SLE patients upon the activation by IL-18, exhibiting significant correlation with the plasma concentrations of Th1 chemokine CXCL10 (all p<0.05). The expression of phospho-JNK in IL-18 activated CD8+ T lymphocytes and the relative % fold increase of the expression of phospho-JNK upon IL-18 activation in B lymphocytes were significantly correlated with SLE disease activity index (both p<0.05). CONCLUSION: The inflammation-mediated activation of JNK and p38 MAPK signaling pathways in T and B lymphocytes can be the underlying intracellular mechanisms causing lymphocyte hyperactivity in SLE.

Decreased expression of T lymphocyte co-stimulatory molecule CD26 on invariant natural killer T cells in systemic lupus erythematosus.

CD26, a T cell co-stimulatory molecule and dipeptidyl peptidase IV for the degradation of interferon-gamma-induced chemokine, participates in multiple immunopathological roles in leukocyte homing and inflammation. Decreased circulating concentration of soluble (s)CD26 in patients with systemic lupus erythematosus (SLE), rheumatoid arthritis and murine model of arthritis and encephalomyelitis have been reported. In the present study, the plasma concentration of sCD26 and chemokines, and cell surface expression of CD26 on monocytes, CD4+T lymphocytes, CD8+T lymphocytes, CD19+B lymphocytes and invariant natural killer T (iNKT) lymphocytes were analyzed using ELISA and flow cytometry, respectively, in 23 SLE patients and 14 sex- and age-matched control subjects. Although there was no significant difference between plasma concentrations of soluble CD26 in SLE patients with controls (p > 0.05), there was significant elevated Th1 chemokines CXCL10 and CXCL9 but not Th2 chemokine CCL2, and down-regulation in iNKT lymphocytes number and cell surface expression of CD26 on CD4+T and iNKT lymphocytes of SLE patients compared with controls (all p < 0.05). Decreased circulating number of iNKT cells and CD26 on iNKT cells can be important for the immunopathogenesis by exacerbating Th1-related inflammation in SLE.