Effect of anti-interferon-γ and anti-interleukin-2 monoclonal antibody treatment on the development of actively and passively induced experimental allergic encephalomyelitis in the SJL/J mouse

SJL/J mice challenged with myelin basic protein (MBP) in complete Freund's adjuvant (CFA) developed only mild chronic-relapsing experimental allergic encephalomyelitis (EAE) with very low incidence. However, treatment of challenged mice with anti-infeferonγ (IFN-γ) monoclonal antibody (mAb) determined severe disease in all cases. Similarly, in passive EAE, the addition of anti-IFN-γ to the in vitro MBP-activated cells at the time of transfer led to significant disease exacerbation in all recipients. The disease enhancing effect was observed only when the mAb was given at the time of active challenge or of passive transfer, but not at later times. Anti-interleukin-2 (IL-2) antibody had only a marginal effect in the active induction, but drastically reduced the manifestations of passive EAE, even when mixed with a disease-enhancing dose of anti-IFN-γ. These findings support the notion that IL-2 is required for disease induction whereas IFN-γ plays a disease-limiting role early in the development of EAE.     

Magnitude of serum and intestinal antibody responses induced by sequential replicating and nonreplicating rotavirus vaccines in gnotobiotic pigs and correlation with protection

A sequential mucosal prime-boost vaccine regimen of oral attenuated (Att) human rotavirus (HRV) priming followed by intranasal (i.n.) boosting with rotavirus protein VP2 and VP6 rotavirus-like particles (2/6-VLPs) has previously been shown to be effective for induction of intestinal antibody-secreting cell (ASC) responses and protection in gnotobiotic pigs. Because serum or fecal antibody titers, but not intestinal ASC responses, can be used as potential markers of protective immunity in clinical vaccine trials, we determined the serum and intestinal antibody responses to this prime-boost rotavirus vaccine regimen and the correlations with protection. Gnotobiotic pigs were vaccinated with one of the two sequential vaccines: AttHRV orally preceding 2/6-VLP (VLP2x) vaccination (AttHRV/VLP2x) or following VLP2x vaccination (VLP2x/AttHRV) given i.n. with a mutant Escherichia coli heat-labile toxin (mLT) as adjuvant. These vaccines were also compared with three i.n. doses of VLP+mLT (VLP3x) and one and three oral doses of AttHRV (AttHRV1x and AttHRV3x, respectively). Before challenge all pigs in the AttHRV/VLP2x group seroconverted to positivity for serum immunoglobulin A (IgA) antibodies. The pigs in this group also had significantly higher (P < 0.05) intestinal IgA antibody titers pre- and postchallenge and IgG antibody titers postchallenge compared to those in the other groups. Statistical analyses of the correlations between serum IgM, IgA, IgG, and virus-neutralizing antibody titers and protection demonstrated that each of these was an indicator of protective immunity induced by the AttHRV3x and the AttHRV/VLP2x regimens. However, only IgA and not IgM or IgG antibody titers in serum were highly correlated (R2 = 0.89; P < 0.001) with the corresponding isotype antibody (IgA) titers in the intestines among all the vaccinated groups, indicating that the IgA antibody titer is probably the most reliable indicator of protection.     

Genotype and phenotype correlation in 103 individuals with 2q37 deletion syndrome reveals incomplete penetrance and supports HDAC4 as the primary genetic contributor

The 2q37 deletion syndrome, also described in the literature as brachydactyly‐mental retardation syndrome (MIM 600430), is caused by deletion or haploinsufficiency of the HDAC4 gene, which encodes the histone deacetylase 4 protein. Although the most commonly described hallmark features of the 2q37 deletion syndrome include brachydactyly type E, developmental delay, obesity, autistic features, and craniofacial or skeletal dysmorphism, a literature review of 101 published cases plus two newly reported individuals indicates that there is a high degree of variability in the presence of some of the features that are considered the most characteristic of the syndrome: overweight and obesity (34%), cognitive‐behavioral issues (79%), dysmorphic craniofacial features (86%), and type E brachydactyly (48%). These features overlap with other neurodevelopmental conditions, including Smith‐Magenis syndrome (SMS), and may be incompletely penetrant or demonstrate variable expressivity, depending on the specific chromosomal anomaly. With the advent of fluorescence in situ hybridization (FISH), array‐based comparative genomic hybridization, and next‐generation DNA sequencing, more detailed molecular diagnoses are possible than in years past, enabling refined characterization of the genotype–phenotype correlation for subjects with 2q37 deletions. In addition, investigations into molecular and gene expression networks are expanding in neurodevelopmental conditions, and we surveyed HDAC4 downstream gene expression by quantitative real‐time polymerase chain reaction, further implicating HDAC4 in its role in the regulation of RAI1. Correlation of clinical data defining the impact on downstream gene expression and the potential clinical associations across neurodevelopment will improve our understanding of these complex conditions and potentially lead to common therapeutic approaches.     

Noradrenergic and opioidergic alterations in neuropathy in different rat strains

The Fischer 344 (F344) rat strain differs from the Lewis strain in the response to neuropathic pain. Recently, we found that F344 rats totally recover from mechanical allodynia induced by chronic constriction injury (CCI) of the sciatic nerve 28 days after surgery whereas Lewis rats are initiating their recovery at this time point. Thus, the use of this neuropathic pain model in these different rat strains constitutes a good strategy to identify possible target genes involved in the development of neuropathic pain. Since differences between Lewis and F344 rats in their response to pain stimuli in acute pain models have been related to differences in the endogenous opioid and noradrenergic systems, we aimed to determine the levels of expression of key genes of both systems in the spinal cord and dorsal root ganglia (DRG) of both strains 28 days after CCI surgery. Real time RT-PCR revealed minimal changes in gene expression in the spinal cord after CCI despite the strain considered, but marked changes in DRG were observed. A significant upregulation of prodynorphin gene expression occurred only in injured DRG of F344 rats, the most resistant strain to neuropathic pain. In addition, we found a significant downregulation of tyrosine hydroxylase and proenkephalin gene expression levels in both strains whereas δ-opioid receptor was found to be significantly downregulated only in injured DRG of Lewis rats although the same trend was observed in F344 rats. The data strongly suggest that dynorphins could be involved in strain differences concerning CCI resistance.     

Noradrenergic neuronal development is impaired by mutation of the proneural HASH-1 gene in congenital central hypoventilation syndrome (Ondine's curse)

Congenital central hypoventilation syndrome (CCHS, Ondine's curse) is a rare disorder of the chemical control of breathing. It is frequently associated with a broad spectrum of dysautonomic symptoms, suggesting the involvement of genes widely expressed in the autonomic nervous system. In particular, the HASH-1-PHOX2A-PHOX2B developmental cascade was proposed as a candidate pathway because it controls the development of neurons with a definitive or transient noradrenergic phenotype, upstream from the RET receptor tyrosine kinase and tyrosine hydroxylase. We recently showed that PHOX2B is the major CCHS locus, whose mutation accounts for 60% of cases. We also studied the proneural HASH-1 gene and identified a heterozygous nucleotide substitution in three CCHS patients. To analyze the functional consequences of HASH-1 mutations, we developed an in vitro model of noradrenergic differentiation in neuronal progenitors derived from the mouse vagal neural crest, reproducing in vitro the HASH-PHOX-RET pathway. All HASH-1 mutant alleles impaired noradrenergic neuronal development, when overexpressed from adenoviral constructs. Thus, HASH-1 mutations may contribute to the CCHS phenotype in rare cases, consistent with the view that the abnormal chemical control of breathing observed in CCHS patients is due to the impairment of noradrenergic neurons during early steps of brainstem development.     

Association between total serum calcium and the A986S polymorphism of the calcium-sensing receptor gene

Serum calcium is under tight physiological control, but it is also a quantitative trait with substantial genetic regulation. Mutations of the CASR gene cause familial hypocalciuric hypercalcemia or autosomal dominant hypoparathyroidism, depending on whether they decrease or increase, respectively, ligand binding to the receptor protein. We described an association between ionized calcium and a common polymorphism (A986S) found in the cytoplasmic tail of this G protein-coupled receptor. We report here on an independent study of 387 healthy young women. Genotyping was performed by allele-specific amplification and serum chemistries were measured by automated clinical assay. Frequencies of SS, AS, and AA genotypes were 6, 107, and 274, respectively, yielding a 986S allele frequency of 15.4%. Mean total serum calcium (Ca(T)) was significantly higher in the SS (9.88 +/- 0.29 mg/dL, P = 0.015) and AS groups (9.45 +/- 0.05 mg/dL, P = 0.002), than in the AA group (9.23 +/- 0.04 mg/dL). In multiple regression modeling, the A986S genotype remained an independently significant predictor of Ca(T) (P < 0.0001) when serum albumin, globulin, inorganic phosphate, and creatinine covariates were included. These data are the first to show significant association between a common polymorphism and concentrations of a serum electrolyte. The A986S polymorphism is also a potential predisposing factor in disorders of bone and mineral metabolism.     

High level expression of apoptosis inhibitor in hepatoma cell line expressing Hepatitis B virus

The serious result of hepatitis B (HBV) virus infection is development of hepatocellular carcinoma (HCC). However, the reason of development of HCC in HBV infected patients is still unclear. Recently, the suppression of cell apoptosis is found to relate with the development of cell carcinogenesis, therefore, the expression of apoptosis inhibitor in the virus related cancer line such as hepatoma cell line HepG2.215 was investigated. There are at least six Human apoptosis inhibitors (IAP) have been identified now. They are cIAP1, cIAP2, XIAP, NAPI, survivin and pIAP. Using gene-assay technology, we have recently compared the expression of IAPs in the HepG2.215 cells that persistently expresses Hepatitis B virus by integrated HBV genome with its parent cell line HepG2. The results suggest that there was obviously increase of cIAP2 and cIAP1 in the HepG2.215 cells versus HepG2 cells. Those observations imply a possibility of long time HBV infection could induce the over-expressing apoptosis inhibitors, furthermore, causing the liver cancer. The high expression of cIAP1 and cIAP2 in HBV expressing cells was confirmed by RT-PCR and Northern blot analysis. However, we did not find the change of NIAP and suvivin in HepG2.215 cells. In contrast, the expression of XIAP was down in the HepG2.215 cells comparing with HepG2 cells. How HBV triggers the over-expression of apoptosis inhibitor is unclear. Transient transfection of HepG2 cells with the plasmids expressing different HBV proteins such as S, M, L, X and core proteins did not give a decisive conclusion. Further study is going on now.     

Duodenal complications in necrotizing pancreatitis: Challenges of an overlooked complication

BackgroundDuodenal complications of necrotizing pancreatitis (NP) are challenging and understudied. We sought to characterize the demographics and clinical course of NP patients with duodenal complications.MethodsSingle institution retrospective review of 687 NP patients treated from 2005 to 2018.ResultsDuodenal complications developed in 40 (6%) patients including fistula in 11 (2%) and stricture in 29 (4%) patients. Patients with duodenal complications had increased computed tomography severity index (CTSI), degree of glandular necrosis, organ failure, infected necrosis, and disease duration. Mortality from NP was increased in patients with duodenal fistula (36%) compared to patients with duodenal stricture (7%) and patients without duodenal complications (9%). Surgical management of duodenal complications was required in 9/11 (82%) patients with fistula and 17/29 (59%) patients with stricture.ConclusionsDuodenal complications occurred in 6% of necrotizing pancreatitis patients. Sixty five percent of patients with duodenal complications required surgical correction. Duodenal fistula was associated with increased mortality.