Intravenous immunoglobulin abrogates dendritic cell differentiation induced by interferon‐α present in serum from patients with systemic lupus erythematosus

Objective

Alterations in the function of dendritic cells (DCs) may explain the systemic autoimmune responses that characterize systemic lupus erythematosus (SLE). Even though several reports have documented the beneficial effect of intravenous immunoglobulin (IVIG) in SLE, the underlying mechanisms of action remain poorly understood. Considering the effect of serum factors, including interferon‐α (IFNα), on the activity of DCs, we investigated the effects of IVIG on the differentiation of DCs mediated by serum from SLE patients.

Methods

DCs were differentiated from peripheral blood monocytes obtained from SLE patients and from healthy blood donors, in the presence of SLE serum. IVIG was used at a concentration of 0.15 mM. A functional assay was performed to assess the inhibitory effect of IVIG on the uptake of nucleosomes by DCs.

Results

IVIG interfered with the differentiation of DCs from SLE patients and healthy donors cultured in the presence of SLE serum. Treatment of DCs with IVIG inhibited the ingestion of nucleosomes by immature DCs, by up to 36%.

Conclusion

The present findings indicate that IVIG, by down‐regulating the IFNα‐mediated differentiation of DCs and by inhibiting uptake of nucleosomes, may exert an essential immunoregulatory effect in SLE patients at the onset of the immune response, at the DC level. Given the critical role of HLA molecules and the costimulatory signals delivered by CD80 and CD86 in optimal antigen presentation and T cell activation, inhibition of expression of HLA and CD80/CD86 on DCs by IVIG offers a plausible explanation for the efficacy of IVIG in SLE and other immune‐mediated inflammatory conditions.

     

The minor pilin PilV provides a conserved adhesion site throughout the antigenically variable meningococcal type IV pilus

Neisseria meningitidis utilizes type IV pili (T4P) to adhere to and colonize host endothelial cells, a process at the heart of meningococcal invasive diseases leading to meningitis and sepsis. T4P are polymers of an antigenically variable major pilin building block, PilE, plus several core minor pilins that initiate pilus assembly and are thought to be located at the pilus tip. Adhesion of N. meningitidis to human endothelial cells requires both PilE and a conserved noncore minor pilin PilV, but the localization of PilV and its precise role in this process remains to be clarified. Here, we show that both PilE and PilV promote adhesion to endothelial vessels in vivo. The substantial adhesion defect observed for pilV mutants suggests it is the main adhesin. Consistent with this observation, superresolution microscopy showed the abundant distribution of PilV throughout the pilus. We determined the crystal structure of PilV and modeled it within the pilus filament. The small size of PilV causes it to be recessed relative to adjacent PilE subunits, which are dominated by a prominent hypervariable loop. Nonetheless, we identified a conserved surface-exposed adhesive loop on PilV by alanine scanning mutagenesis. Critically, antibodies directed against PilV inhibit N. meningitidis colonization of human skin grafts. These findings explain how N. meningitidis T4P undergo antigenic variation to evade the humoral immune response while maintaining their adhesive function and establish the potential of this highly conserved minor pilin as a vaccine and therapeutic target for the prevention and treatment of N. meningitidis infections.

The human-restricted bacterial pathogen Neisseria meningitidis is a leading cause of meningitis and sepsis worldwide and represents a significant global public health threat (1, 2). N. meningitidis is carried asymptomatically in the protective mucus layer of the throat for 5 to 25% of the population (3–5). In some cases, N. meningitidis disseminates into the bloodstream, an environment to which this bacterium is remarkably well adapted. Meningococci possess a polysaccharide capsule that protects them against complement deposition plus several membrane associated factors that are important for survival, including factor H binding protein and iron uptake systems (6). Critical to N. meningitidis survival in the bloodstream are the type IV pili (T4P), which mediate vascular colonization; nonpiliated meningococci are rapidly cleared from the blood (7–9). T4P are long filamentous appendages displayed peritrichously on the bacterium. The major pilin protein, PilE, is the primary building block of the pilus. This and other surface-displayed N. meningitidis proteins undergo antigenic variation, allowing this pathogen to evade a protective immune response (10–12).T4P are responsible for acute colonization of human blood vessels and are thus essential in establishing invasive meningococcal diseases (7, 9, 13, 14). T4P are helical polymers of the major pilin assembled by the T4P machinery (15, 16). The conserved N terminus of the major pilin is a hydrophobic α-helix that tethers the C-terminal globular domain in the inner membrane prior to pilus assembly and forms a helical array in the core of the intact pilus, displaying the globular domain on the filament surface. Pilus assembly is initiated by a cluster of pilin-like proteins called minor pilins (17–20). These “core” minor pilins are thought to localize to the pilus tip. The major pilin, PilE, is highly conserved in amino acid sequence and structure between N. meningitidis and the urogenital pathogen Neisseria gonorrhoeae with the exception of a hypervariable β-hairpin near the C terminus that is prominent on the pilus surface (21–24). In N. meningitis, PilE has been shown to bind to sialylated N-glycans on the human endothelial cell receptor CD147 (also called EMMPRIN or Basigin) (25) and the β2-adrenergic receptor (26), two membrane proteins that form a heterotrimeric complex with cytoplasmic α-actinin 4 (27).The pathogenic Neisseria possess a set of core minor pilins, PilH (FimT), PilI (PilV), PilJ (PilW), PilK, and PilX, that are encoded within a single gene cluster and prime pilus assembly (28), plus “noncore” minor pilins PilV and ComP, which are encoded elsewhere on the genome. ComP shares the canonical T4P–pilin structure of the major pilin, PilE, with the N-terminal α-helix and C-terminal globular domain (29). ComP is involved in natural transformation of exogenous DNA (30). PilV, which is highly conserved in N. meningitidis isolates (31), participates in adhesion and signaling in host cells (13, 26, 32–34). In N. meningitidis, PilV, like PilE, directly interacts with CD147 and the β2-adrenergic receptor, suggesting that PilV colocalizes with PilE within the T4P filament (13, 26). However, another report concluded that PilV functions exclusively from within the periplasm, fine-tuning pilus surface display to regulate its interactions with host cells (35).The T4P–receptor interaction represents a key step in N. meningitidis adhesion and colonization of endothelial cells in peripheral and brain vasculature and is thus an attractive target for preventive and therapeutic approaches to tackle meningococcal infection. Interfering with piliation prevents N. meningitidis colonization of human endothelial cells and vasculature (36) and improves sepsis outcome in a mouse model grafted with human skin (9). Although both PilE and PilV are involved in adhesion, PilE exhibits considerable amino acid sequence variability in its exposed hypervariable region. This variability contributes to Neisseria immune escape. In contrast, PilV is highly conserved and has been shown to be immunogenic in humans (37). Thus, PilV may prove to be a more promising target than PilE for blocking endothelial cell adhesion. A molecular understanding of this minor pilin with respect to its structure, localization within the pilus, and interactions with host receptors will be valuable in assessing its potential as a therapeutic target. Here, we report the atomic structure of PilV and superresolution microscopy images showing that it is incorporated throughout the N. meningitidis T4P. We identify residues involved in adhesion to host cells and map these onto the PilV structure, modeled within the cryoelectron microscopy (cryoEM)-derived pilus filament structure. Finally, we show that anti-PilV antibodies inhibit meningococcal adhesion in vivo. These data provide insights into PilV-mediated adhesion and suggest that blocking its adherence functions may inhibit N. meningitidis vascular colonization and pathogenesis.     

Breastfeeding initiation or duration and longitudinal patterns of infections up to 2 years and skin rash and respiratory symptoms up to 8 years in the EDEN mother–child cohort

This paper aimed to examine the effect of breastfeeding on longitudinal patterns of common infections up to 2 years and respiratory symptoms up to 8 years. To assess the incidence and reoccurrence of infections and allergic symptoms in the first years of life among 1,603 children from the EDEN mother–child cohort, distinct longitudinal patterns of infectious diseases as well as skin rash and respiratory symptoms were identified by group‐based trajectory modelling. To characterize infections, we considered the parent‐reported number of cold/nasopharyngitis and diarrhoea from birth to 12 months and otitis and bronchitis/bronchiolitis from birth to 2 years. To characterize allergy‐related symptoms, we considered the parent‐reported occurrence of wheezing and skin rash from 8 months to 8 years and asthma from 2 to 8 years. Then associations between breastfeeding and these longitudinal patterns were assessed through adjusted multinomial logistic regression. Compared with never‐breastfed infants, ever‐breastfed infants were at a lower risk of diarrhoea events in early infancy as well as infrequent events of bronchitis/bronchiolitis throughout infancy. Only predominant breastfeeding duration was related to frequent events of bronchitis/bronchiolitis and infrequent events of otitis. We found no significant protective effect of breastfeeding on longitudinal patterns of cold/nasopharyngitis, skin rash, or respiratory symptoms. For an infant population with a short breastfeeding duration, on average, our study confirmed a protective effect of breastfeeding on diarrhoea events in early infancy, infrequent bronchitis/bronchiolitis and, to a lesser extent, infrequent otitis events up to 2 years but not on other infections, skin rash, or respiratory symptoms4.     

MEK in cancer and cancer therapy

The mitogen-activated extracellular signal-regulated kinase (MEK) pathway is one of the best-characterized kinase cascades in cancer cell biology. It is triggered by either growth factors or activating mutations of major oncogenic proteins in this pathway, the most common being Ras and Raf. Deregulation of this pathway is frequently observed and plays a central role in the carcinogenesis and maintenance of several cancers, including melanoma, pancreatic, lung, colorectal, and breast cancers. Targeting these kinases offers promise of novel therapies. MEK inhibitors (MEKi) are currently under evaluation in clinical trials and many have shown activity. In this review, we comprehensively examine the role of the MEK pathway in carcinogenesis and its therapeutic potential in cancer patients, with a focus on MEKi. We describe the clinical perspectives of MEKi in the two main models of Ras–ERK driven tumors, BRAF-mutant (“addicted” to the pathway) and KRAS-mutant (non-“addicted”). We also highlight the known mechanisms of resistance to MEKi and emerging strategies to overcome it.     

The pyridinyl-6 position of WAY-100635 as a site for radiofluorination--effect on 5-HT1A receptor radioligand behavior in vivo.

PURPOSE: We aimed to evaluate radiofluorination at the pyridinyl-6 position of the selective 5-HT(1A) receptor antagonist, WAY-100635 [N-(2-(1-(4-(2-methoxyphenyl)piperazinyl)ethyl))-N-(2-pyridinyl)cyclohexanecarboxamide)], on 5-HT(1A) receptor radioligand behavior in vivo. PROCEDURES: The pyridinyl-6 [(18)F]fluoro derivative of WAY-100635 ([(18)F]6FPWAY) was obtained by direct nucleophilic substitution with [(18)F]fluoride ion in a bromo precursor. After intravenous injection of [(18)F]6FPWAY into Cynomolgus monkey, the uptake of radioactivity into brain regions was assessed with positron emission tomography (PET) and blood samples analyzed by high performance liquid chromatography (HPLC) for parent radioligand and radioactive metabolites. The experiment was repeated after pretreatment of the monkey with a dose of WAY-100635 that blocks brain 5-HT(1A) receptors. RESULTS: After intravenous injection of [(18)F]6FPWAY into Cynomolgus monkey, the uptake of radioactivity into whole brain reached 4.33% of injected dose at 7.5 min. Uptake was highest in 5-HT(1A) receptor-rich regions. Pretreatment with WAY-100635 reduced uptake in these regions to near the levels in receptor-devoid cerebellum. [(18)F]6FPWAY was rapidly metabolized in vivo, as evidenced by the rapid appearance of radioactive metabolites in plasma. CONCLUSION: [(18)F]6FPWAY is selective and moderately useful for imaging brain 5-HT(1A) receptors in vivo. The pyridinyl-6 position is resistant to defluorination and may be an attractive site for the (18)F-labeling of 6FPWAY analogs that resist hydrolysis.