Th17 responses to pneumococcus in blood and adenoidal cells in children.

Pneumococcal infections cause a large global health burden, and the search for serotype-independent vaccines continues. Existing conjugate vaccines reduce nasopharyngeal colonization by target serotypes. Such mucosal effects of novel antigens may similarly be important. CD4+ Th17 cell-dependent, antibody-independent reductions in colonization and enhanced clearance have been described in mice. Here we describe the evaluation of T helper type 17 (Th17) cytokine responses to candidate pneumococcal protein vaccine antigens in human cell culture, using adenoidal and peripheral blood mononuclear cells. Optimal detection of interleukin (IL)-17A was at day 7, and of IL-22 at day 11, in these primary cell cultures. Removal of CD45RO+ memory T cells abolished these responses. Age-associated increases in magnitude of responses were evident for IL-17A, but not IL-22, in adenoidal cells. There was a strong correlation between individual IL-17A and IL-22 responses after pneumococcal antigen stimulation (P < 0·015). Intracellular cytokine staining following phorbol myristate acetate (PMA)/ionomycin stimulation demonstrated that  > 30% CD4+ T cells positive for IL-22 express the innate markers γδT cell receptor and/or CD56, with much lower proportions for IL-17A+ cells (P < 0·001). Responses to several vaccine candidate antigens were observed but were consistently absent, particularly in blood, to PhtD (P < 0·0001), an antigen recently shown not to impact colonization in a clinical trial of a PhtD-containing conjugate vaccine in infants. The data presented and approach discussed have the potential to assist in the identification of novel vaccine antigens aimed at reducing pneumococcal carriage and transmission, thus improving the design of empirical clinical trials.

Protein-Derived Acetaminophen-Cysteine Can Be Detected After Repeated Supratherapeutic Ingestion of Acetaminophen in the Absence of Hepatotoxicity.

UNLABELLED: Generation of protein-derived acetaminophen-cysteine (APAP-CYS) is reported after ingestion of large and therapeutic dosages of acetaminophen in healthy and in liver-damaged patients. The incidence of protein-derived APAP-CYS adducts in repeated supratherapeutic dosages of APAP is not known. METHODS: for 12 months, a standardized and comprehensive questionnaire was used to interview every consecutive patient at a pain management clinic. Patients found to ingest more than 4 g of APAP per day for a minimum of 14 consecutive days at the time of the encounter were invited to have blood drawn for hepatic transaminases and APAP-CYS adduct levels. Twelve subjects out of 990 interviewees met inclusion criteria. Ten of the 12 had measurable protein-derived APAP-CYS, none had evidence of liver injury. Patients that ingest repeated supratherapeutic amounts of APAP over several weeks may generate APAP-CYS protein adducts in the absence of hepatic injury.

Voriconazole is safe and effective as prophylaxis for early and late fungal infections following allogeneic hematopoietic stem cell transplantation.

Seventy-two patients undergoing allogeneic transplantation were treated with voriconazole (VOR) as antifungal prophylaxis starting from day -2 of transplantation and continuing until withdrawal of immunosuppression. Patients were assessed for safety and the incidence of definite, probable, or possible fungal infection throughout transplantation was evaluated. VOR was well tolerated. Only 14% of patients required interruption of VOR therapy because of toxicity: liver toxicity (8%), cardiac Q-T interval prolongation (1%), or other side effects (5%). In the early post-transplant period (<120 days), only 2 patients developed invasive fungal infection: 1 mucormycosis infection and 1 disseminated Aspergillus infection. In the late post-transplant period (>120 days), no patients developed probable or definite fungal infection while receiving VOR. No Candida infections were seen in either period. These data suggest that fungal prophylaxis with VOR following allogeneic transplantation is safe and effective.

Proteomics identifies enhanced expression of stefin A in neonatal murine skin compared with adults: functional implications.

BACKGROUND: Skin develops through a process of epidermal proliferation, maturation, and remodelling of the epidermis and dermis. This period also involves the maturation of the skin immune system, such that antigen applied though the skin of a neonatal mouse always results in immunosuppression, whereas in adults, immunity will occur. OBJECTIVES: Using proteomics, to identify proteins uniquely involved in the development of the skin and skin immune system. METHODS: Proteins were extracted from whole skin of mice aged 4 and 21 days, and separated using two-dimensional electrophoresis. RESULTS: Of the 25 proteins that were sequenced by peptide mass fingerprinting with matrix-assisted laser desorption/ionization-time of flight-mass spectrometry, three were known markers of keratinocyte differentiation and proliferation. These were cyclophilin A, epidermal fatty acid binding protein 5 and stefin A. Of interest were the two isoforms of stefin A, an intracellular protease inhibitor, found in neonatal skin. The strong expression of stefin A in neonates was confirmed by immunohistochemical analysis, suggesting an important role in the development of the epidermis. Additionally, Western blotting identified two larger isoforms in adult skin, revealing a change in the stefin A during development. CONCLUSIONS: We propose that stefin A is involved in development of the skin, that development of the skin and of immune function is linked, and that stefin A has an important function in neonatal skin and potentially the neonatal immune response.

Carcinogen-modified dendritic cells induce immunosuppression by incomplete T-cell activation resulting from impaired antigen uptake and reduced CD86 expression.

Exposure of the skin to environmental stimuli, such as chemical or physical carcinogens, modifies the local skin environment, including depletion of epidermal Langerhans' cells (LC). Any subsequent exposure of the LC-depleted skin to antigen results in the generation of antigen-specific tolerance. In this study we evaluated the antigen-bearing cells in the draining lymph nodes by capitalizing on the fluorescent nature of the contact sensitizer, fluorescein isothiocyanate (FITC). When FITC was applied to the skin of normal mice, two distinct populations of antigen-bearing cells were identified in the draining lymph nodes. They were classified as either FITChi or FITClo on the basis of their fluorescence intensity and thus the amount of antigen they internalized. Only FITClo cells were detected in the lymph nodes draining FITC-treated murine skin that had been depleted of epidermal LC by prior treatment with the complete carcinogen 9,10-dimethyl 1,2-benzanthracene (DMBA). Functional analysis of these cells revealed that the FITChi cells, but not the FITClo cells, induced antigen-specific T-cell proliferation. Further analysis of the FITClo cells from the DMBA-treated mice demonstrated that these cells had reduced levels of CD80 expression, had substantially reduced levels of CD86 expression and performed poorly as co-stimulator cells in an anti-CD3-mediated proliferative assay. Nonetheless these cells still induced early signs of T-cell activation and interleukin-12 production. Consequently the FITClo cells migrating from the LC-depleted skin, through a combination of reduced antigen presentation and reduced co-stimulatory activity, induced a state of unresponsiveness or anergy in the responder T cells in a similar manner to that observed when antigen presentation occurs in the absence of co-stimulation. We propose that these unresponsive, or anergic cells, account for the antigen-specific tolerance observed in these experiments.

Vitreoretinal findings similar to retinopathy of prematurity in infants with compound heterozygous protein S deficiency.

OBJECTIVE: To present previously undescribed vitreoretinal findings similar to severe retinopathy of prematurity (ROP) in two siblings (daughter and son) with a thrombophilic disorder, compound heterozygous protein S (PS) deficiency. DESIGN: Family genotype study and literature review. PARTICIPANTS: Two unrelated heterozygous PS-deficient parents and their two children with compound heterozygous PS deficiency were studied. The gestational age and birth weight of the daughter were 40 weeks and 3200 g, respectively, and those of the son were 34 weeks and 2150 g, respectively. Three other neonates with homozygous or compound heterozygous PS deficiency and ophthalmologic findings were identified in the literature. INTERVENTION: The daughter underwent lensectomy-vitrectomy at 48 weeks adjusted age bilaterally. The son underwent therapy developed for severe ROP: laser therapy of the peripheral avascular retina at 39 weeks adjusted age, and bilateral lensectomy-vitrectomy with membrane peel of intravitreous proliferation from the optic disc at 42 weeks adjusted age. MAIN OUTCOME MEASURES: The main clinical outcome measures were retinal appearance and functional vision. Genotypes of the family members were determined. RESULTS: One of the four eyes retained functional vision. A normal-appearing posterior retina, normal scotopic and photopic flash electroretinograms, and a normal flash visual-evoked response were documented from the left eye of the son at 62 weeks adjusted age. The other three eyes had inoperable retinal detachments and no functional vision. The mother had type I PS deficiency and the father had type II PS deficiency. Compound heterozygous PS deficiency was confirmed in both children. CONCLUSION: In both children, normal vasculogenesis was interrupted. At 39 weeks adjusted age, the retinal examination of the son revealed extraretinal fibrovascular proliferation at the optic disc (reactivation of the hyaloid system) and in the peripheral retina (interruption of inner retinal vascularization). Patients with homozygous or compound heterozygous PS deficiency may present as infants with severe ROP. The authors' experience suggests that appropriately timed surgical procedures, which are efficacious for ROP, can preserve vision in infants with thrombophilic disorders.

Necrotizing fasciitis of the pharynx following adenotonsillectomy.

Necrotizing fasciitis is a rare clinical entity in the head and neck region. We report a case of necrotizing fasciitis following adenotonsillectomy in a previously healthy 2-year-old girl. The child presented in a septic state with impending airway compromise. Computed tomography (CT) showed massive soft tissue widening with air in the retropharyngeal, parapharyngeal and retromandibular spaces. Intraoperative exploration showed necrosis of the posterior pharyngeal wall from the skull base to the cricoid, with extension into the parapharyngeal and retropharyngeal spaces. Cultures from the debrided tissues grew two aerobes and three anaerobes. Management involved airway support, surgical debridement, broad spectrum antibiotic coverage and nutritional support. The patient ultimately developed nasopharyngeal and oropharyngeal stenosis requiring tracheostomy and gastrostomy tube placement. This case report highlights an extremely rare complication of adenotonsillectomy.

Reduction of respiratory syncytial virus (RSV) in tracheal aspirates in intubated infants by use of humanized monoclonal antibody to RSV F protein.

Thirty-five children <2 years of age mechanically ventilated for respiratory syncytial virus (RSV) infection were randomized to receive an intravenous infusion of 15 mg/kg MEDI-493 or placebo. RSV concentration was measured in tracheal secretions by plaque assay before and at 24-h intervals after treatment. The reduction in tracheal RSV concentration from day 0 to day 1 (-1.7+/-0.28 vs. -0. 6+/-0.21 log10 pfu/mL; P=.004) and from day 0 to day 2 (-2.5+/-0.26 vs. -1.0+/-0.41 log10 pfu/mL; P=.012) was significantly greater in the MEDI-493 group than in the placebo group. RSV concentration in nasal aspirates did not differ significantly between the groups. No significant differences were observed in the tracheal aspirate white blood cell count, or myeloperoxidase or eosinophilic cationic protein concentration, or in measures of disease severity between the groups. Thus, treatment with 15 mg/kg MEDI-493 intravenously was well-tolerated and significantly reduced RSV concentration in tracheal aspirates of children with respiratory failure due to RSV.

Timing of the occurrence of pulmonary embolism in trauma patients.

OBJECTIVE: To determine how soon after trauma pulmonary embolism (PE) occurs and if there is an association between the duration of this interval and mortality. DESIGN: Retrospective case series. PATIENTS: All patients admitted to our trauma service with established PE based on high probability findings on ventilation perfusion scan, positive results on a pulmonary arteriogram, or autopsy from July 1, 1990, to September 30, 1995. MAIN OUTCOME MEASURE: Time interval between injury and PE. SETTING: Level I university trauma center. RESULTS: Of 18,255 trauma patients identified, 63 met our criteria for PE (30 using a pulmonary arteriogram; 26, a ventilation perfusion scan; and 7, autopsy). Four patients (6%) had a documented PE on day 1 following injury. Mortality was not correlated with the interval between injury and PE. Of the 63 patients, 58 (92%) had 1 or more established risk factors for thromboembolism. The ratio of PaO2 to fraction of inspired oxygen was the only factor predictive of mortality (P = .02, logistic regression analysis). CONCLUSIONS: Pulmonary embolism occurs in the immediate period following injury. Aggressive workup in patients with signs consistent with PE should be instituted promptly. Trauma patients who have at least 1 risk factor for thromboembolism should receive prophylaxis as soon after injury as possible.

Long-term form identification vision after early, closed, lensectomy-vitrectomy for stage 5 retinopathy of prematurity.

PURPOSE: Form identification vision after early, closed, lensectomy-vitrectomy for retinopathy of prematurity (ROP) stage 5 open funnel retinal detachment is reported from a database that included 45 eyes of 27 infants. The focus of this report is the verbal responses at a mean age of 7.0 years for nine nonamblyopic (preferred) eyes of nine preterm infants with minimal developmental delay (good central nervous system function). METHODS: All 45 eyes underwent initial cryotherapy for threshold ROP to the avascular retina to decrease the angiogenic stimulus (mean postconceptual age = 34.8 weeks) and subsequently underwent multiple cryotherapy sessions to the avascular retina and shunt with scleral buckling to decrease retinal traction (mean postconceptual age = 38.0 weeks). When tractional retinal detachment occurred with an open funnel, each eye underwent an early, closed, lensectomy-vitrectomy (mean postconceptual age = 45.7 weeks). The 34 eyes with a successful anatomic result were fitted with contact lenses as soon as possible after surgery. RESULTS: The nine nonamblyopic eyes of nine preterm infants with minimal developmental delay had the following visual acuities using Allen figures or Snellen test types: one eye 20/80, one eye 20/200, two eyes 20/400, three eyes 20/800, and two eyes 20/ 1600. CONCLUSION: These nine eyes support the thesis that form identification vision can be obtained by early vitrectomy for ROP stage 5 open funnel retinal detachments.

Primary ocular malignant lymphoma associated with the acquired immune deficiency syndrome.

A 42-year-old man who was human immunodeficiency virus (HIV)-positive complained of floaters in his right eye, which had existed for 1 week, followed by loss of central vision. Results of ophthalmoscopic examination disclosed confluent yellowish-white retinochoroidal infiltrates with perivascular sheathing, which were more prominent superiorly in the right eye. Approximately 10 small, white, intraretinal and choroidal lesions were observed in the nasal periphery of the left eye. Results of cytologic examination of the vitreous of the right eye showed neoplastic cells characteristic of large cell type malignant lymphoma. Shortly thereafter, cytologic examination of the cerebrospinal fluid also showed large cell malignant lymphoma. Magnetic resonance imaging (MRI) showed a mass involving the left cerebellar hemisphere. After bilateral whole-eye radiation therapy, there was complete resolution of the lymphomatous retinochoroidal infiltrates in both eyes. The ophthalmologic and neurologic manifestations of acquired immune deficiency syndrome (AIDS) are discussed. The authors believe this is the first report of ocular malignant lymphoma occurring in a patient with AIDS.

In vitro and in vivo studies with purified recombinant human interleukin 5.

The functional activities of highly purified recombinant human IL 5 (hIL 5) have been characterized on a number of cell types in vitro and in BALB/c mice in vivo. In vitro, hIL 5 could induce the differentiation of eosinophils from precursors in both human and mouse bone marrow with approximately the same efficiency. A mouse IL 5/3-dependent B cell line, LyH7.B13, was found to proliferate in response to hIL 5 but not human interleukin 1 (IL 1), interleukin 2 (IL 2), interleukin 3 (IL 3), interleukin 4 (IL 4), interleukin 6 (IL 6), interferon-gamma (IFN-gamma), or granulocyte macrophage-colony stimulating factor (GM-CSF) and was at least 10-fold more sensitive than BCL1 mouse lymphoma cells. We have successfully used this cell line to demonstrate the production of IL 5 by human T cell clones. In marked contrast to its effects on murine B cell lines, hIL 5 had no demonstrable activity on CD23 expression, anti-mu costimulated proliferation or IgM, IgG, or IgE production by tonsillar B cells and did not influence such responses triggered by IL 4. BALB/c mice injected with hIL 5 for 7 consecutive days were shown to develop an eosinophilia comparable to that induced by infection with the parasite Mesocestoid corti.

Modification of protein synthesis initiation factors and the shut-off of host protein synthesis in adenovirus-infected cells.

A substantial body of data, largely derived from study of cell extracts, indicates that protein synthesis in adenovirus-infected cells requires VA RNAI at late times of infection to prevent the activation of a protein kinase known as DAI, and the consequent phosphorylation of the alpha-subunit of initiation factor eIF-2. To verify this conclusion, we have measured the steady-state levels of eIF-2 alpha phosphorylation in cells infected with wild-type virus (Ad2) and a mutant that produces no VA RNAI (Ad5dl331). Consistent with the proposed mechanism, the alpha-subunit was very highly phosphorylated (approximately 90%) at late times of infection with Ad5dl331. Surprisingly, eIF-2 alpha phosphorylation also increased (to approximately 30%) at late times of infection with Ad2, suggesting that VA RNA and DAI might be involved in the selective translation of viral mRNA and the shut-off of host cell protein synthesis during the late phase. In agreement with this model, host protein synthesis shut-off is defective in cells expressing low levels of DAI.

A mechanism for the control of protein synthesis by adenovirus VA RNAI.

In the absence of VA RNAI, protein synthesis in adenovirus-infected HeLa cells fails because of defective initiation. Earlier work showed that the defect results from phosphorylation of the initiation factor elF-2 on its alpha subunit. We have identified the protein kinase responsible as the dsRNA-activated inhibitor of protein synthesis (DAI). DAI is present in uninfected state. It is activated in cells infected with the adenovirus mutant Ad5 dl331, which produces no VA RNAI, but not in cells infected with wild-type virus. Activation occurs during the late phase of infection with the mutant virus, and the activator appears to be dsRNA produced by symmetrical transcription of the viral genome. VA RNAI antagonizes the activation of DAI by dsRNA, but it cannot inhibit the activity of DAI once activated. We propose a mechanism for VA RNAI action based on its partially double-stranded nature.

Pigmented free-floating vitreous cysts in two young adults. Electron microscopic observations.

Pigmented free-floating vitreous cysts were observed in two young adults. In both patients, the cyst was in the visual axis; however, the size and extent of pigmentation of the cyst wall compromised the visual acuity only in case 1. In this case, the vitreous cyst was aspirated through the pars plana and studied by light and electron microscopy. Histopathologically, the cyst was lined by a heavily pigmented layer of cuboidal cells intermixed with sheets of nonpigmented cells forming papillae. Ultrastructurally, the pigmented cells contained predominantly large, mature melanosomes (0.9-2.2 micron). Scattered immature melanosomes with a scarcity of mitochondria and other cytoplasmic organelles were present. Additionally, the cells were invested by a thin polarized basement membrane and displayed apical microvilli. Numerous microvillous processes were noted under the plasmalemmae and between adjacent cells. The results of the light and ultrastructural studies provide support for the hypothesis that the cyst in case 1 originated from the pigment epithelium. The possibility of a traumatic etiology is proposed for these pigmented vitreous cysts. If significant visual impairment is present, surgical removal of the cyst through a pars plana approach can be safely performed as in our case 1.

Retinal detachment following late posterior capsulotomy.

When we compared the incidence of retinal detachment in 27 eyes after extracapsular cataract surgery and late posterior capsulotomy with the incidence of general aphakic detachment, we found no relationship between the onset of retinal detachment and the numbers, types, or locations of breaks. The interval between cataract extraction and discission had a significant effect on recovery of visual acuity. Final visual acuities of 20/400 or better were attained in 22 of 23 eyes in which discission followed cataract extraction by one year or more but in only one of four eyes in which the procedures were separated by less than one year. Delaying discission for more than 12 months may reduce the extent of retinal detachment and produce the best functional results. Detachments that occurred less than one year after discission were more extensive but achieved better functional results than those that occurred later.

Mouse skin tumor-initiating activity of 5-, 7-, and 12-methyl- and fluorine-substituted benz[a]anthracenes.

As an approach for elucidation of structure activity relationships that underlie the exceptionally large difference in carcinogenic activity between benz[a]anthracene and 7,12-dimethylbenz[a]anthracene (7,12-DMBA), 11 methyl- and/or fluorine-substituted benz[a]anthracenes were evaluated for tumor-initiating activity on mouse skin. Outbred CD-1 and outbred Sencar mice received a single topical application of the hydrocarbons followed by twice weekly applications of the tumor promoter 12-O-tetradecanoylphorbol 13-acetate for 16-26 weeks. 7,12-DMBA was almost two orders of magnitude more active as a tumor-initiator than 7- and 12-methylbenz[a]anthracene. Methyl substitution at the 7- and 7,12-positions of benz[a]anthracene was significantly more effective in the enhancement of tumorigenic activity than fluorine substitution at these positions. Although 7-fluorobenz[a]anthracene, 12-fluorobenz[a]anthracene, and 7,12-difluorobenz[a]anthracene had only 0.15, 0.26, and less than 0.005 times the tumor-initiating activity of their respective methyl-substituted derivatives, they were severalfold more active than benz[a]anthracene. 7-Fluorobenz[a]anthracene was slightly less active than 12-fluorobenz[a]anthracene, whereas 7-methylbenz[a]anthracene was about twofold more than 12-methylbenz[a]anthracene. For 7,12-di-substituted benz[a]anthracenes, 7-methyl-12-fluorobenz[a]anthracene was more than twice as tumorigenic as 7-fluoro-12-methylbenz[a]anthracene, but each was individually more active than 7-methylbenz[a]anthracene and 12-methylbenz[a]anthracene, respectively. Both fluorinated compounds were much less active than 7,12-DMBA. Substitution of fluorine or methyl at the 5-position of 7-methylbenz[a]anthracene and substitution of fluorine at the 5-position of 12-methylbenz[a]anthracene dramatically reduced their tumorigenic activity.

Visual acuity after surgery for retinal detachment with macular involvement.

The records for a group of 2,054 patients who had undergone surgery fur rhegmatogenous retinal detachment with macular involvement were analyzed to ascertain the relative influence of the following five specific factors on postoperative visual acuity: (1) patient age, (2) degree of preoperative macular elevation, (3) duration of preoperative macular detachment, (4) extent of retinal detachment, and (5) drainage of subretinal fluid. Increasing patient age, increasing preoperative macular elevation, increasing duration of macular detachment, and increasing extent of retinal detachment were all found to be associated in general with decreasing postoperative visual acuity. Drainage, or nondrainage, or subretinal fluid appeared to be unassociated with postoperative visual acuity.

Management of the fellow eyes of patients with rhegmatogenous retinal detachment.

The experience of the senior author over the past 18 years in the management of fellow eyes of 3,036 patients with rhegmatogenous detachment is reported. The study group is composed of 2,221 patients with no history of detachment surgery or prophylactic treatment in the functional fellow eye. Of 816 eyes with retinal breaks, lattice degeneration, or retinoschisis-the types of pathology most conducive to detachment-70% were treated with xenon-arc photocoagulation or cryoapplication. Detachment occurred subsequently in 5.4% of the eyes treated with photocoagulation, 2% of the eyes treated with cryoapplications, and 8.2% of the eyes with major pathology that were not treated. Essentially normal fellow eyes with no major pathology and, consequently no treatment, had a subsequent detachment rate of 5.3%. The authors conclude that prophylactic treatment via transconjunctival cryoapplication is safe and effective and recommend treatment of all fellow eyes with lattice degeneration and breaks.

Aphakic macular oedema following prosthetic lens implantation.

Fluorescein angiography of the iris was performed on patients with plastic lens implants with cystoid oedema of the macula, and the nature of the vascular changes was compared with controls provided by patients who did not have macular disease. Densitometry was used to quantitate leakage of fluorescein dye into the anterior chamber in residual angiograms 5 minutes after injection. Cystoid oedema was associated with marked increase in vascular permeability of the iris in the patients with implants, and in aphakics without implants, but to a lesser degree. Although it cannot be concluded that implants themselves cause either the increase in permeability or the onset of macular oedema, the presence of such implants must be considered an additional hazard in an eye which is already at risk. Iris angiography can be used as a diagnostic aid in patients in whom macular oedema is suspected. A persistent increase in permeability associated with a reduction in visual acuity should act as a warning of further visual loss and of eventual cystoid macular degeneration.