Ocular and oral grading of mucous membrane pemphigoid.

BACKGROUND: A variety of methods have been described for grading ocular mucous membrane pemphigoid (MMP), each with their own limitations. In contrast, there are no reported grading systems for involvement of the oral mucosa. We wished to evaluate two ocular (one established and one proposed) and an oral mucosal grading system for MMP. METHODS: Patients with MMP were assessed by three ophthalmologists and two oral medicine physicians. Ocular disease was graded using the system described by Rowsey and a proposed system based on measurement of vertical depth and horizontal width measured from the bulbar conjunctival aspect. Oral assessment used a 'mucosal disease severity score' originally described for lichen planus, in which 17 areas of the mouth are scored for involvement, together with a pain score. Levels of agreement were evaluated using Fleiss' Kappa Statistic (k). RESULTS: Forty-four patients with MMP encompassing mild to severe disease were included. Good levels of agreement were observed between observers for both vertical (k:0.86) (upper 95% CI: 1.03 mm) and horizontal (k:0.80) (upper 95% CI: 3.01 mm) involvement for the proposed ocular system and the Rowsey system (k: 0.83) (upper 95% confidence interval: 3.19 mm). There was a high coefficient of determination (R(2)) between the ocular grading systems (0.81, p < 0.01). Oral grading showed excellent levels of agreement (k: 0.71) between observers. There was no significant association between the severity of oral and ocular disease using described grading systems. CONCLUSIONS: The proposed grading systems for both oral and ocular involvement in MMP are easy to use, and show good agreement between observers. The proposed ocular system correlates well with a currently used system, and overcomes some of the difficulties encountered with existing systems. For the individual patient, changes greater than 1.5 mm (vertical) and 3 mm (horizontal) are significant. This may increase our ability to detect change or disease progression. Although the risk of ocular involvement in patients with only oral involvement has been demonstrated, the severity of oral and ocular disease are not well-correlated, due in part to an absence of an ocular disease activity score.

Prospective study of surgical outcomes and bleb morphology using indocyanine green as a surgical dye in trabeculectomy with mitomycin C.

BACKGROUND: To investigate the effect of adding indocyanine green to mitomycin C in augmented trabeculectomy. DESIGN: A prospective, non-comparative interventional case series. PARTICIPANTS: A total of 37 eyes of 37 patients followed up for 1 year. METHODS: A solution containing 12.5 mg/mL of indocyanine green was added to mitomycin C, resulting in an mitomycin C concentration of 0.2-0.4 mg/mL, which was applied to bare sclera and Tenon's capsule for 3 min during trabeculectomy. MAIN OUTCOME MEASURES: Visual acuity, intraocular pressure, bleb morphology, Moorfields Bleb Grading System scores and complications. RESULTS: Indocyanine green could be visualized on clinical examination for all eyes on the first postoperative day. Mean intraocular pressure decreased from 22.9 ± 6.2 mmHg to 12.1 ± 4.4 mmHg postoperatively (P < 0.001) at 1 year. Thirty-four eyes (91.9%) achieved an intraocular pressure of less than 21 mmHg at final visit without additional topical intraocular pressure-lowering medications. Three eyes (8.1%) developed bleb failure and required Baerveldt device implantation. There were no cases of blebitis or late bleb leak. No adverse effects attributable to indocyanine green could be identified postoperatively. CONCLUSION: The addition of indocyanine green during trabeculectomy improves the visibility of antimetabolites intraoperatively and allows for the estimation of antimetabolite treatment area intraoperatively and postoperatively. It appears to have no adverse effect on surgical outcomes and complication rates, while improving safety of antimetabolite use.

Resistance to and recovery from lethal influenza virus infection in B lymphocyte-deficient mice.

In the adaptive immune response to most viruses, both the cellular and humoral arms of the immune system play complementary roles in eliminating virus and virus-infected cells and in promoting recovery. To evaluate the relative contribution of CD4+ and CD8+ effector T lymphocytes in virus clearance and recovery, we have examined the host response to lethal type A influenza virus infection in B lymphocyte-deficient mice with a targeted disruption in the immunoglobulin mu heavy chain. Our results indicate that naive B cell-deficient mice have a 50- 100-fold greater susceptibility to lethal type A influenza virus infection than do wild type mice. However, after priming with sublethal doses of influenza, immune B cell-deficient animals show an enhanced resistance to lethal virus infection. This finding indicates that an antibody-independent immune-mediated antiviral mechanism accounts for the increased resistance to lethal virus challenge. To assess the contribution of influenza-specific CD4+ and CD8+ effector T cells in this process, defined clonal populations of influenza-specific CD4+ and CD8+ effector T cells were adoptively transferred into lethally infected B cell-deficient mice. Cloned CD8+ effectors efficiently promoted recovery from lethal infection, whereas cloned CD4+ T cells conferred only partial protection. These results suggest that memory T lymphocytes can act independently of a humoral immune response in order to confer resistance to influenza infection in immune individuals. The potential implications of these results for vaccination against human influenza infection are discussed.

Influenza virus-specific CD4+ T helper type 2 T lymphocytes do not promote recovery from experimental virus infection.

T lymphocytes play a primary role in recovery from viral infections and in antiviral immunity. Although viral-specific CD8+ and CD4+ T cells have been shown to be able to lyse virally infected targets in vitro and promote recovery from lethal infection in vivo, the role of CD4+ T lymphocytes and their mechanism(s) of action in viral immunity are not well understood. The ability to further dissect the role that CD4+ T cells play in the immune response to a number of pathogens has been greatly enhanced by evidence for more extensive heterogeneity among the CD4+ T lymphocytes. To further examine the role of CD4+ T cells in the immune response to influenza infection, we have generated influenza virus-specific CD4+ T cell clones from influenza-primed BALB/c mice with differential cytokine secretion profiles that are defined as T helper type 1 (Th1) clones by the production of interleukin 2 (IL-2) and interferon gamma (IFN-gamma), or as Th2 clones by the production of IL-4, IL-5, and IL-10. Our studies have revealed that Th1 clones are cytolytic in vitro and protective against lethal challenge with virus in vivo, whereas Th2 clones are noncytolytic and not protective. Upon further evaluation of these clonal populations we have shown that not only are the Th2 clones nonprotective, but that pulmonary pathology is exacerbated as compared with control mice as evidenced by delayed viral clearance and massive pulmonary eosinophilia. These data suggest that virus-specific CD4+ T cells of the Th2 subset may not play a primary role in virus clearance and recovery and may lead to immune mediated potentiation of injury.

Response to influenza infection in mice with a targeted disruption in the interferon gamma gene.

Interferon gamma (IFN-gamma) is a pleiotropic cytokine secreted by T lymphocytes and natural killer (NK) cells and has been noted to be a first line of host defense in the control of viral infections. To examine further the role of this cytokine in the control of viral infections, mice with a targeted mutation in the IFN-gamma gene were infected with influenza virus, and the in vivo antibody and cell-mediated immune response to viral infection were examined. In addition, cell lines and clones were derived from the immunized animals and the in vitro cytokine production and cytotoxic T lymphocyte (CTL) response were analyzed. The absence of IFN-gamma led to increased production of influenza-specific IgG1, IL-4, and IL-5 as compared to wild-type littermate control animals. In contrast, there was no difference noted in the development of an effective CTL response between IFN-gamma-deficient and wild-type animals. In this model of experimental influenza infection, IFN-gamma is not necessary for the development of an effective humoral or cellular immune response to challenge with this respiratory virus.

Antibody recognition of an immunogenic influenza hemagglutinin-human leukocyte antigen class II complex.

The A/Japan/57 influenza hemagglutin (HA) peptide HA 128-145, when bound by human histocompatibility leukocyte antigen-DRw11 cells, is recognized by the human CD4+ T cell clone V1. A rabbit antiserum has been raised against HA 128-145 which recognizes not only the free peptide, but also the HA 128-145/DRw11 complex on a solid matrix, in solution, or on the surface of viable cells. The detection of these complexes on viable cells was shown to be class II specific, DRw11 restricted, and commensurate with the level of DRw11 expression. The identity of DRw11 as the cell surface molecule binding HA 128-145 was confirmed by immunoprecipitation, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and tryptic peptide mapping. Using this antiserum HA 128-145/DRw11 complexes could be detected on the cell surface as soon as 30 min after the peptide was added, and increased up to 24 h. Dissociation kinetics showed these complexes were long-lived, with a half-life of approximately 14 h. This anti-HA peptide antiserum represents the first direct means of studying antigenic peptide-human leukocyte antigen class II complexes on the surface of living cells without the addition of a non-amino acid moiety to the peptide. The properties of this antiserum thus provide the potential to study naturally processed antigenic peptides as well as the mechanism of processing itself in a physiologically relevant system.

Development of a highly detailed virtual model eye.

Using a three-dimensional modeling computer program, the authors created a three-dimensional, highly detailed computer-generated model of a normal eye to enhance the teaching of basic ophthalmic anatomy. The resulting model is a uniquely detailed virtual eye that can illustrate ocular anatomy clearly but realistically at many levels of magnification. Eye anatomy can be demonstrated from any viewpoint, with a choice of which structures are visible. The generation of video animations is also possible. The potential of this model for illustration of ocular disease will be the objective of further testing with medical students.

Serological studies of peptostreptococci using an indirect fluorescent antibody test.

Members of the genus Peptostreptococcus are frequent isolates from periodontal lesions. A study was undertaken for determination of the serological relationships of oral and non-oral strains of several species of this genus. Eighty-nine strains of peptostreptococci representing seven species were tested by means of an indirect fluorescent antibody technique (IFA), with rabbit antisera to Peptostreptococcus anaerobius ATCC 27337, Peptostreptococcus micros VPI 2618-A, and Peptostreptococcus productus ATCC 27340. Each antiserum showed positive fluorescence when reacted with homologous cells. When anti-P. micros VPI 2618-A serum was added to suspensions of P. micros ATCC 33270 and clinical isolates of P. micros in the IFA, positive fluorescence was observed to a titer of 1:64 with 26 out of 29 strains. Positive fluorescence was also seen when P. anaerobius VPI 5737 and clinical isolates of P. anaerobius were tested with anti-P. anaerobius ATCC 27337 serum; a titer of at least 1:64 was observed in 10 out of 14 strains. These results support the presence of two serological groups of P. anaerobius, rabbit anti-P. anaerobius ATCC 27337 serum reacting with Group II organisms. No interspecies cross-reactivity was observed except with four strains of Peptostreptococcus magnus, which reacted with several antisera as well as with normal rabbit serum and the saline controls. These results indicate that rabbit antisera and the use of an IFA are useful in the identification of P. anaerobius and P. micros from oral lesions.

Use of Staurenghi lens angiography in the management of posterior uveitis.

PURPOSE: Retinal vasculitis is an important component of posterior uveitis. It may be difficult to detect either clinically or with conventional angiography. We assessed the role of Staurenghi lens angiography (SLA) in the diagnosis and management of patients with posterior uveitis. METHODS: A total of 26 patients attending the St Paul's Eye Unit uveitis clinic with posterior uveitis underwent wide-angle (150 degree) retinal angiography with the Heidelberg Retina Angiograph 2 Scanning Laser Ophthalmoscope using the Staurenghi SLO 230 contact lens. We determined the percentage of patients in whom this imaging modality assisted in diagnosing and quantifying the extent of vasculitis, planning treatment or monitoring disease activity. RESULTS: Staurenghi lens angiography assisted in diagnosing and quantifying the extent of vasculitis in 62% of patients, and in planning laser photocoagulation or immunosuppression titration in 62% of patients and enhanced disease monitoring in 35% of patients. In 31% of cases, SLA revealed vasculitis that was not evident clinically. CONCLUSION: Staurenghi lens angiography is a valuable tool in the management of patients with posterior uveitis and can be used for the diagnosis, treatment and follow-up of these patients.

Corneal confocal microscopy in Darier disease.

PURPOSE: We report ocular findings in a patient with Darier disease. In particular, we discuss the in vivo confocal images. METHOD: A retrospective noninterventional case report. RESULTS: Confocal microscopy revealed collections of intercellular material, separating the cells of the patient's basal corneal epithelium. Discrete foci of this material were scattered throughout the central corneal basal epithelium in both eyes. These deposits had a similar reflectance as corneal nerves, were 3-5-microm thick, and of note in places conformed to the cell borders. CONCLUSIONS: The findings of abnormal intercellular collections at the basal epithelial layer of the cornea correlate with the previously reported histological findings. They are consistent with the abnormal keratocyte-to-keratocyte adhesions that are described both in the skin and cornea of Darier disease. The abnormalities we describe occurred in the absence of other ocular signs of Darier disease. These findings on confocal microscopy may help with the diagnosis and monitoring of disease progression.

Serological reactions of the genus Peptostreptococcus.

White male New Zealand rabbits were immunized with soluble antigen preparations (SP) of the following gram-positive anaerobic cocci: Peptostreptococcus anaerobius ATCC 27337 and VPI 5737; P. micros VPI 2618; Streptococcus morbillorum ATCC 27527; P. parvulus VPI 5229; and P. productus ATCC 27340. SP were reacted with homologous and heterologous rabbit antisera in immunodiffusion, immunoelectrophoresis, indirect fluorescent-antibody, and tanned-cell passive hemagglutination tests. Even though each antiserum reacted strongly with its homologous SP, no interspecies reactivity was observed except between P. productus antisera and P. parvulus SP by the passive hemagglutination test. Antisera prepared to both strains of P. anaerobius reacted with the other strain in all serological tests.

Extractable antigen shared by Peptostreptococcus anaerobius strains.

Extracts from several species of gram-positive cocci were prepared by a modification of the Rantz-Randall autoclave method and tested for reactions with rabbit anti-Peptostreptococcus anaerobius (ATCC 27337 and VPI 5737) sera in a capillary precipitin test. Antigen preparations from two reference strains of P. anaerobius (ATCC 27337 and VPI 5737) and six clinical isolates of P. anaerobius reacted with the P. anaerobius antisera. These extracts formed a line of identity by immunodiffusion and displayed at least one precipitin line by immunoelectrophoresis. Absorption of the antisera with either the autoclaved extract or a 10% whole-cell suspension from each of the eight P. anaerobius strains removed the precipitin line(s) observed during immunodiffusion and immunoelectrophoresis. Extracts prepared to other species of Peptococcus, Peptostreptococcus, and Streptococcus did not react with the P. anaerobius antisera in a capillary precipitin test. In addition antisera to Lancefield groups A to G did not react with the extracts from the eight P. anaerobius strains. Preliminary chemical analysis of the extracts from the eight strains showed that they contained approximately 0.2 mg of carbohydrate per ml and 3.6 mg of protein per ml. The rabbit anti-P. anaerobius sera used in this study detected a common antigen(s) shared by strains of P. anaerobius, but did not react with autoclave extracts prepared from other species of gram-positive cocci. This extractable antigen could be used in a capillary precipitin test to rapidly identify P. anaerobius strains isolated in the clinical microbiology laboratory.

IL-2 induces Fas ligand/Fas (CD95L/CD95) cytotoxicity in CD8+ and CD4+ T lymphocyte clones.

IL-2 is a T cell growth factor that has pleiotropic functions in T cell differentiation, induction of lymphokine-activated killer cells, and regulation of immune responses. In studying TCR triggering of perforin or Fas ligand (FasL)/Fas (CD95 ligand/CD95) cytotoxicity in our influenza-specific T cell clones, we found that IL-2 can also induce FasL/Fas cytotoxicity. IL-2 induces FasL/Fas cytotoxicity in our CD8+ and CD4+ Th1 clones, but not in our CD4+ Th2 clones. IL-2 induction of cytolytic activity occurs when the CD8+ T cells are refractory to IL-2-induced proliferation. This killing is Ag independent, MHC unrestricted, and blocked by Fas.Fc fusion protein. IL-2 induces FasL/Fas cytotoxicity in a dose-dependent manner, but does not induce high levels of FasL expression as detected by flow cytometry. TCR triggered FasL/Fas cytotoxicity is detectable in CD8+ and Th1 clones by 3 h and peaks at 6 h; high levels of killing are maintained for at least 24 h. Similarly, IL-2 induces FasL/Fas killing in CD8+ and Th1 clones within 3 h of stimulation and maintains high levels for at least 24 h. TCR-triggered FasL/Fas killing is inhibited by emetine and cyclosporin A, whereas IL-2-induced FasL/Fas killing is inhibited by emetine, but not by cyclosporin A. These results demonstrate a second mechanism to induce FasL/Fas cytotoxicity in CD8+ and Th1 clones and may explain IL-2 induction of Ag-independent MHC-unrestricted lymphokine-activated killer cell activity.

Infections due to vancomycin-resistant Enterococcus faecium resistant to linezolid.

Linezolid is a new oxazolidinone antibiotic used to treat infections caused by vancomycin-resistant enterococci (VRE). In early clinical trials, emergence of resistance occurred rarely. We report clinical details and antibiotic susceptibility from five patients treated with linezolid for VRE infections who had resistant organisms isolated during therapy. Four were transplant patients receiving protracted courses of the drug; three cases were associated with treatment failure. One of 45 linezolid-treated patients developed resistance during therapy. Susceptibility testing should be done in all cases on starting therapy.

Antibody reactive with Peptostreptococcus micros in the sera of patients with periodontal disease.

Subgingival plaque samples were cultured for the isolation of Peptostreptococcus micros from individuals with and without chronic periodontitis. Humoral antibody from each person reacted with P. micros in an indirect fluorescent-antibody test, but not all of the sera reacted in a passive hemolysis test. No correlations were observed between the presence of antibody reactive with P. micros and the isolation of P. micros or the gingival health of the individual.

Recognition of the influenza hemagglutinin by class II MHC-restricted T lymphocytes and antibodies. I. Site definition and implications for antigen presentation and T lymphocyte recognition.

We have identified the site encompassing residues 126-145 on the A/Japan/57 influenza hemagglutinin molecule that is recognized in association with HLA-DRw11 by a clonal population of human, influenza specific, CD4+ cytolytic T lymphocytes. The critical core sequence of the T cell determinant spans hemagglutinin residues 129-140 and overlaps a putative antibody binding site. Hemagglutinins of influenza field strains that are not recognized by the T cell clones contain sequence alterations within the 129-140 target site of the CD4+ T cells. Functional analyses, with synthetic peptides, of the contribution of each of the residues within the sequence toward the capacity of the antigenic fragment to associate with both the restriction element and the TCR revealed a continuous linear array of residues necessary for MHC binding and/or Ag receptor engagement. At least one residue, the lysine at position 134, was shown to be critical for both DRw11 association and TCR recognition. The significance of these findings for recognition of glycoproteins by human CD4+ T cells is discussed.