Mitsuaki Kimura

Correlation of house dust mite-specific lymphocyte proliferation with IL-5 production, eosinophilia, and the severity of symptoms in infants with atopic dermatitis.

Because house dust mite (HDM)-specific IgE antibody (IgE-RAST) is usually not detectable in infants with atopic dermatitis (AD), HDM has not been regarded as the cause of infantile AD. The level of HDM-specific lymphocyte proliferation (expressed as stimulation index measured by flow cytometry [SIF]), however, was found to be markedly elevated in AD infants. This suggests that the sensitization of T cells to HDM extract occurs even in infancy. Moreover, the level of HDM-SIF is correlated closely with the severity of infantile AD, suggesting that HDM is a major cause of this disease not only in adults and children but also in infants. Although the level of HDM-SIF did not correlate with the level of HDM-specific IgE-RAST in infants with AD, it did intimately correlate with the absolute number of peripheral blood eosinophils. Because T lymphocytes are known to secrete some cytokines, such as IL-5, that enhance the proliferation of eosinophils, we measured the IL-5 production by peripheral blood mononuclear cells in infants with AD on stimulation with HDM extract. The amount of IL-5 production is significantly higher in infants with AD, as well as in children with AD, than that found in nonatopic control subjects. Moreover, the level of IL-5 production is correlated closely with the level of HDM-SIF in infants with AD. Taken together, these results suggest that HDMs play an important role in the development of infantile AD by inducing IL-5 production from HDM-specific T lymphocytes.

IL-4 production by PBMCs on stimulation with mite allergen is correlated with the level of serum IgE antibody against mite in children with bronchial asthma.

BACKGROUND Although IL-4, IL-13, and IFN-gamma are known to affect IgE synthesis, it remains unclear which one plays the most important role in in vivo IgE synthesis in atopic patients. OBJECTIVE The aim of this study is to clarify the difference in importance among these cytokines in up-regulation of IgE synthesis in atopic patients. METHODS We measured IL-4, IL-13, and IFN-gamma production by PBMCs on stimulation with house dust mite (HDM) in 23 children, 3 to 15 years old, with bronchial asthma (BA) and analyzed the correlation with HDM-specific IgE antibody levels expressed as HDM IgE radioallergosorbent test (RAST) results. RESULTS The production of IL-4 and IL-13 by PBMCs on stimulation with HDM was significantly higher in children with BA than in nonatopic control subjects (IL-4, 752.9 +/- 365.9 vs 312.3 +/- 230.0 fg/mL, P <.001; IL-13, 21.9 pg/ml [<12.0-77.6] vs <12.0, P <.01). IL-4 production showed a close positive correlation with HDM IgE RAST (r = 0.71, P <.001), which was distinctly stronger than that between IL-13 production and HDM IgE RAST (r = 0.46, P <.05). IFN-gamma production was neither different between children with BA and nonatopic control subjects (7. 24 [1.54-33.90] pg/mL vs 11.2 [1.66-75.9] pg/mL) nor correlated with HDM IgE RAST levels. Essentially the same result was obtained by stimulation of PBMCs with a purified HDM major allergen Der f 1. CONCLUSION IL-4 is likely to be the most important cytokine in up-regulation of in vivo IgE synthesis against HDM in children with BA.

Unique profile of IL-4 and IFN-γ production by peripheral blood mononuclear cells in infants with atopic dermatitis

Although the level of house dust mite (HDM)-specific lymphocyte proliferation is high in infants with atopic dermatitis (AD), the level of HDM-specific IgE antibody (HDM-IgE-RAST) is usually very low or negative. To elucidate the cause of the deficient HDM-specific IgE antibody formation in infants with AD, we examined the profile of IL-4 and IFN-gamma production by HDM-stimulated PBMCs. The amount of IL-4 production was higher in infants with AD and in children with AD (3 to 15 years) than in the nonatopic control subjects. Although the amount of IFN-gamma production in children with AD was lower than that found in nonatopic children, it was higher in infants with AD than in nonatopic infants. This result suggests that HDM-specific helper T lymphocytes in infants with AD have not yet differentiated into TH2 but rather stayed at the stage of TH0. The level of IgE-RAST for egg white (EW) is already elevated in infants with AD. The amount of IL-4 produced by EW-stimulated PBMCs was comparable to that produced by HDM-stimulated PBMCs in infants with AD. However, the amount of IFN-gamma produced by EW-stimulated PBMCs was distinctly lower than that produced by HDM-stimulated PBMCs in infants with AD. Although there was no correlation between the amount of IL-4 production by HDM-stimulated PBMCs and the level of HDM-IgE-RAST in infants with AD, the amount of IL-4 production by EW-stimulated PBMCs was closely correlated with the level of EW-IgE-RAST. These results suggest that it is not the lack of IL-4 but rather a relative increase in IFN-gamma production by HDM-specific helper T lymphocytes that causes the deficiency of HDM-specific IgE-antibody synthesis in infants with AD.

Skin infiltration of CD56bright CD16- natural killer cells in a case of X-SCID with Omenn syndrome-like manifestations

We observed a patient with X‐linked severe combined immunodeficiency (X‐SCID) with Omenn syndrome‐like manifestations. X‐linked inheritance, absence of CD132 expression and impaired response to interleukin‐2 (IL‐2) indicated that the case is typical of X‐SCID due to γc defect. However, this case was unusual in that circulating natural killer (NK) cells were increased and nearly half of these NK cells exhibited the CD56bright CD16− phenotype. A missense mutation was found within exon 5 of the IL2RG gene. The identical mutation was detected within NK, CD4+ T and B cells. Engraftment of maternally derived NK cells or gene reversion was ruled out. The erythroderma‐like skin lesion was characterized by infiltration of the dermis by CD56bright NK cells admixed with CD1a+ dendritic cells (DC). Expression of mRNA for inflammatory cytokines was significantly enhanced within the skin. This may be the first human case to demonstrate that close cell‐to‐cell contact between DC and NK cells provides an effective alternative pathway for NK cell differentiation/activation in vivo.

Usefulness of lymphocyte stimulation test for the diagnosis of intestinal cow's milk allergy in infants.

Background: Bottle-fed infants sometimes develop intestinal cow’s milk allergy (ICMA). Because cow’s milk-specific IgE antibody (CM-IgE) levels are normal, the lymphocyte stimulation test (LST) has been proposed as an alternative diagnostic test for ICMA. The present study evaluated the diagnostic value of LST in a large number of patients with ICMA in Japan. Methods: Ninety-six infants who developed intestinal symptoms after ingestion of cow’s milk formula and showed remission of symptoms after elimination of this food were enrolled as patients with probable ICMA. Seventy-two subjects with normal CM-IgE levels and a positive result in an oral food challenge test (OFCT) for cow’s milk formula were diagnosed with ICMA. Another 10 infants with normal CM-IgE levels and a negative OFCT result were diagnosed with nonspecific intestinal symptoms (NIS). The status of cell-mediated immunity against cow’s milk proteins was estimated by LST for ĸ-casein. Results: In the 72 patients with ICMA (38 boys and 34 girls), the median age at onset was 9 days. Sixty-two of 72 (86.1%) patients with ICMA tested positive in the LST for ĸ-casein. In contrast, only 2 of the 10 NIS infants tested positive. The incidence of a positive LST result was significantly higher in the ICMA group than in the NIS group (p < 0.0001). The area under the receiver-operating characteristic curve for this test was as high as 0.856. Conclusions: This study strongly suggests that the LST for ĸ-casein is a useful diagnostic test for ICMA.

Development of the Capacity of Peripheral Blood Mononuclear Cells to Produce IL-4, IL-5 and IFN-γ upon Stimulation with House Dust Mite in Children with Atopic Dermatitis

Background: Imbalances of IL-4 and IFN-γ production are widely known to increase IgE synthesis in allergic individuals, and IL-5 is known to play a critical role in the pathogenesis of various allergic diseases. However, little is known about how Th cells specific to house dust mite (HDM) develop the capacity to produce these cytokines in children with atopic dermatitis (AD). Objective: This study aims to clarify when HDM-specific Th cells develop the capacity to produce IL-4, IL-5 and IFN-γ in children with AD. Methods: The production of IL-4, IL-5 and IFN-γ by peripheral blood mononuclear cells (PBMCs) upon stimulation with HDM was measured in 91 children with AD and 37 control subjects. The changes in the cytokine production with age were analyzed transectionally and longitudinally. Results: IL-4 production by HDM-stimulated PBMCs in children with AD was already increased before age 1. Thereafter, it continuously rose until reaching a near-maximum level at age 2. Growth-related changes in the production of IL-5 resembled those in the production of IL-4 and peaked at age 1. The production of these cytokines was very low in control subjects up to age 2 and then gradually increased, albeit never above the levels measured in AD children. The production of IFN-γ in children with AD reached a near-maximum level during the first year of life and diminished after age 3. Although IFN-γ production in controls was lower than that in AD children during infancy, it continuously rose even after age 3 and surpassed the levels measured in AD children. The level of serum HDM-specific IgE antibody began to increase after age 1 following the rise of IL-4 production. Essentially the same relationship among IL-4, IFN-γ and IgE synthesis was seen in a longitudinal study of 6 AD infants, in whom HDM-specific IgE was initially negative but later turned positive. Conclusions: These findings suggest that the capacity of HDM-specific Th cells to produce IL-4, IL-5 and IFN-γ develops and effectively matures during the first 3 years of life in children with AD.

Phosphatase and tensin homolog (PTEN) mutation can cause activated phosphatidylinositol 3-kinase δ syndrome–like immunodeficiency

BACKGROUND Activated phosphatidylinositol 3-kinase δ syndrome (APDS) is a recently discovered primary immunodeficiency disease (PID). Excess phosphatidylinositol 3-kinase (PI3K) activity linked to mutations in 2 PI3K genes, PIK3CD and PIK3R1, causes APDS through hyperphosphorylation of AKT, mammalian target of rapamycin (mTOR), and S6. OBJECTIVE This study aimed to identify novel genes responsible for APDS. METHODS Whole-exome sequencing was performed in Japanese patients with PIDs. Immunophenotype was assessed through flow cytometry. Hyperphosphorylation of AKT, mTOR, and S6 in lymphocytes was examined through immunoblotting, flow cytometry, and multiplex assays. RESULTS We identified heterozygous mutations of phosphatase and tensin homolog (PTEN) in patients with PIDs. Immunoblotting and quantitative PCR analyses indicated that PTEN expression was decreased in these patients. Patients with PTEN mutations and those with PIK3CD mutations, including a novel E525A mutation, were further analyzed. The clinical symptoms and immunologic defects of patients with PTEN mutations, including lymphocytic AKT, mTOR, and S6 hyperphosphorylation, resemble those of patients with APDS. Because PTEN is known to suppress the PI3K pathway, it is likely that defective PTEN results in activation of the PI3K pathway. CONCLUSION PTEN loss-of-function mutations can cause APDS-like immunodeficiency because of aberrant PI3K pathway activation in lymphocytes.

Differences in cytokine production by peripheral blood mononuclear cells (PBMC) between patients with atopic dermatitis and bronchial asthma

It is widely accepted that type 2 helper T (Th2) lymphocytes play a crucial role in the pathogenesis of atopic dermatitis (AD) as well as bronchial asthma (BA). We measured the amounts of IL‐5 and interferon‐gamma (IFN‐γ) produced by PBMC upon stimulation with house dust mite (HDM) or Candida albicans (CA) in 17 children (3–15 years) with AD, and compared these values with those of 16 children with BA. Although IL‐5 production by PBMC upon stimulation with HDM in patients with AD was significantly higher than that in 13 non‐atopic controls (geometric mean = 23.4 pg/ml versus 5.9 pg/ml, P < 0.05), it was significantly lower than that in patients with BA (177.8 pg/ml, P < 0.001). The amount of IL‐5 produced by PBMC upon stimulation with CA was also significantly lower in patients with AD than in those with BA (7.2 pg/ml versus 100.0 pg/ml, P < 0.001). The production of IFN‐γ by PBMC stimulated with HDM or CA was also significantly lower in patients with AD than in those with BA (HDM 4.3 pg/ml versus 12.6 pg/ml, P < 0.05; CA 6.5 pg/ml versus 60.3 pg/ml, P < 0.001). Consequently the ratio of IL‐5 to IFN‐γ production was high not only in patients with BA but also in those with AD. These findings suggest that there are some differences in the regulation of in vivo cytokine production between patients with AD and those with BA, although a Th2‐dominant profile is common to both.

Increased CRP level and fever in infants with FPIES in Japan

Increased C‐reactive protein (CRP) and fever are observed in some infants with food protein‐induced enterocolitis syndrome (FPIES) in Japan, but the reproducibility of these findings has not yet been confirmed on oral food challenge (OFC).

Increased C‐reactive protein and fever in Japanese infants with food protein‐induced enterocolitis syndrome

Increased C‐reactive protein (CRP) and fever are observed in some infants with food protein‐induced enterocolitis syndrome (FPIES) in Japan, but the reproducibility of these findings has not yet been confirmed on oral food challenge (OFC).