Arachnoid granulations of the temporal bone: a histologic study of dural and osseous penetration.

HYPOTHESIS: Arachnoid granulations (AG) are more prevalent along the middle fossa surface of the temporal bone, where they produce larger bony defects than those occurring on the posterior surface. BACKGROUND: Dural and bony defects formed by AGs are proposed to lead to spontaneous meningoencephaloceles and cerebrospinal fluid otorrhea. They most commonly occur at the tegmen and in individuals older than 40 years. METHODS: Vertically sectioned temporal bones were evaluated using light microscopy to determine AG histology, distribution, and morphometry and to determine the prevalence of AG penetration in the donor population. RESULTS: AGs were observed to penetrate the dura mater and make direct contact with cortical surfaces in 12.7% of donors in the Johns Hopkins Temporal Bone Collection. AGs occurred at middle fossa sites 13% more frequently than at posterior fossa sites. At middle fossa sites AGs produced significantly larger bony openings and were more likely to be associated with herniating brain tissue. Donors with AGs were significantly older, and all were in the late 30s or older. CONCLUSION: Erosion of the temporal bone by AGs is not a rare occurrence in the population and becomes increasingly prevalent with age. It is estimated that 14 in 1,000 donors were at greatest risk of eventual cerebrospinal fluid leakage at the tegmen. The age and anatomic distribution described in this study strengthens the notion that AG penetration plays a role in the pathophysiology of spontaneous cerebrospinal fluid leaks and meningoencephaloceles of the temporal bone.

Inflammatory cytokine expression on the ocular surface in the Botulium toxin B induced murine dry eye model.

PURPOSE: Inflammation plays an important role in dry eye syndrome. In this study, inflammatory cytokine expression on the ocular surface in the Botulium toxin B (BTX-B) induced mouse dry eye model was investigated. METHODS: CBA/J mice received an injection of saline or 20 milliunits (mU) of BTX-B into the lacrimal gland. Tear production and corneal fluorescein staining were evaluated in all groups before injection and at 3 time points after. The pro-inflammatory cytokines macrophage inhibitory factor (MIF), interleukin-1beta (IL-1 beta), tumor necrosis factor-alpha (TNF- alpha) and interleukin-6 (IL-6) in conjunctival and corneal epithelium were evaluated by real time quantitative PCR and immunohistochemistry. RESULTS: BTX-B injected mice showed significantly decreased aqueous tear production and increased corneal fluorescein staining at the 1 week and 2 week time points compared with normal control and saline-injected mice. The BTX-B injected mice mRNA expression levels of TNF-alpha and IL-1beta from conjunctival and corneal epithelial cells increased significantly at two early time points comparing with that of normal and saline injected mice, but IL-1beta returned to normal levels at the 4 week time point. Saline injected mice showed no difference in mRNA expression of TNF-alpha, IL-1beta, MIF, and IL-6 on the ocular surface tissue at all time points. Immunohistochemistry confirmed these findings. CONCLUSIONS: BTX-B induced mouse model showed decreased aqueous tear production, increased corneal fluorescein staining, and TNF-alpha and IL-1beta increased expression on the ocular surface within one month. The patterns seen appeared to mimic those in humans with non-Sjögren's syndrome keratoconjunctivitis sicca (NS-KCS).

Assessment of Pentalogy of Cantrell using 3D Multidetector Computed Tomography.

A 7-month-old white female with Pentalogy of Cantrell was imaged using 64 slice multidetector computed tomography (MDCT) with 3D mapping to better determine the extent of cardiac, thoracic, and abdominal malformations. Complimentary to ultrasound, the use of 3D 64 slice MDCT can facilitate effective diagnosis and treatment planning in cases of Pentalogy of Cantrell.

Corneal wound healing is modulated by topical application of amniotic fluid in an ex vivo organ culture model.

The purpose of this study was to evaluate the effects of topical human amniotic fluid (HAF) and equine amniotic fluid (EAF) on corneal reepithelialization and stromal wound healing. New Zealand white rabbit corneas (n=52) were placed in an ex vivo air-interface organ culture. An 8.5mm-diameter mark in the center of the cornea was produced with a hand trephine to select the area for epithelial scraping. A number 15 surgical blade was used to remove the epithelial layer within the demarcated area in a standardized fashion. The corneas were assigned to one of four treatment groups (n=8): fetal bovine serum (FBS), HAF, EAF, and a control group that was exposed to phosphate buffer solution (PBS). Corneal epithelial defects were imaged every 8 h for 72 h after the application of a 30 microl drop of 0.015% fluorescein. Five corneas of each treatment group were used for histology, proliferation, and apoptosis assay at 72 h after the epithelial defect was created. There was no significant difference in the mean rate of closure of the corneal epithelial defect between FBS treated corneas and controls (P>0.06). The mean epithelial defect area (MEDA) was significantly smaller in the EAF group as compared to control corneas at 24 h (P=0.016), 40 h (P=0.032), 64 h (P=0.008) and 72 h (P=0.007) following epithelial scrape. The MEDA in the HAF group was significantly smaller at 16 h (P=0.008), 64 h (P=0.0072), and 72 h (P=0.016) compared to the control group. The MEDA in the HAF and EAF groups was smaller at all time points as compared to the FBS group, but the difference was not significant. At histology, the mean keratocyte density was significantly higher in the anterior stroma in the HAF (P<0.001) and EAF groups (P=0.001) as compared to control group. The number of BrdU positive keratocytes was significantly higher in the superficial and deep stromal sub-areas in the HAF group as compared to control (P<0.001 and P=0.002, respectively). EAF and FBS treated corneas also showed a higher number of BrdU positive cells compared to control, but this difference was not significant. Finally, we did not observe any difference in the amount of TUNEL positive keratocytes among the different groups. Our data indicates that the topical application of HAF and EAF is associated with accelerated reepithelialization in this cornea organ culture model. Similarly, corneal keratocyte density appears to be less affected after epithelial injury using this treatment.

Periocular triamcinolone enhances intraocular gene expression after delivery by adenovirus.

PURPOSE: A noninvasive imaging technique was used for serial assessment of gene expression after intraocular gene transfer. Bioluminescence after intravitreous administration of an adenovirus vector containing the firefly luciferase gene was measured serially and noninvasively. The optical signal was then used as a bioassay to determine whether periocular immune modulation affects intraocular transgene expression. METHODS: Sixty-two, 8-week-old, male BALB/c mice were used. The correlation of optical signal intensity was determined by tissue luciferase level after injecting 30 mice with one of three intravitreous doses of Ad-Luc-GFP (10(8), 5 x 10(8), or 10(9) particles in 1 microL). Ocular bioluminescence was measured at days 2, 5, 8, and 14. The bioluminescence was then directly compared with measured tissue luciferase levels. The remaining 32 mice were divided into two groups. One group (n = 16), was injected with periocular corticosteroid (400 mug in 10 microL). Two days later, Ad-Luc-GFP was administered by intravitreous injection (10(9) particles in 1 microL). The remaining mice (n = 16) were injected with the same dose of intravitreous Ad-Luc-GFP without corticosteroid pretreatment. Ocular bioluminescence was then assessed longitudinally on days 2, 4, 6, 8, 11, 14, 22, and 30 after intravitreous injection in n = 10 mice per group. The optical signal intensity in each group was compared over the study period. The remaining 12 mice (n = 6, each group) were used to assess histologic differences between the two groups. RESULTS: In vivo measurement of ocular bioluminescence was well correlated with tissue luciferase levels (Spearman's correlation, r = 0.969, P < 0.001). Periocular TA injection markedly decreased the acute inflammatory reaction associated with intravitreous Ad-Luc-GFP and was associated with a significant increase in the duration of peak luciferase expression as well as the total period of luciferase expression. CONCLUSIONS: A significant enhancement of intraocular transgene expression is associated with periocular pretreatment with corticosteroid. Histologic evidence of immune cell reduction in ocular tissues in corticosteroid-treated eyes implies a local immune response. Periocular treatment with corticosteroids may enhance adenovirus-mediated gene expression in the eye.

Use of molecular modeling and site-directed mutagenesis to define the structural basis for the immune response to carbohydrate xenoantigens.

BACKGROUND: Natural antibodies directed at carbohydrates reject porcine xenografts. They are initially expressed in germline configuration and are encoded by a small number of structurally-related germline progenitors. The transplantation of genetically-modified pig organs prevents hyperacute rejection, but delayed graft rejection still occurs, partly due to humoral responses. IgVH genes encoding induced xenoantibodies are predominantly, not exclusively, derived from germline progenitors in the VH3 family. We have previously identified the immunoglobulin heavy chain genes encoding VH3 xenoantibodies in patients and primates. In this manuscript, we complete the structural analysis of induced xenoantibodies by identifying the IgVH genes encoding the small proportion of VH4 xenoantibodies and the germline progenitors encoding xenoantibody light chains. This information has been used to define the xenoantibody/carbohydrate binding site using computer-simulated modeling. RESULTS: The VH4-59 gene encodes antibodies in the VH4 family that are induced in human patients mounting active xenoantibody responses. The light chain of xenoantibodies is encoded by DPK5 and HSIGKV134. The structural information obtained by sequencing analysis was used to create computer-simulated models. Key contact sites for xenoantibody/carbohydrate interaction for VH3 family xenoantibodies include amino acids in sites 31, 33, 50, 57, 58 and the CDR3 region of the IgVH gene. Site-directed mutagenesis indicates that mutations in predicted contact sites alter binding to carbohydrate xenoantigens. Computer-simulated modeling suggests that the CDR3 region directly influences binding. CONCLUSION: Xenoantibodies induced during early and delayed xenograft responses are predominantly encoded by genes in the VH3 family, with a small proportion encoded by VH4 germline progenitors. This restricted group can be identified by the unique canonical structure of the light chain, heavy chain and CDR3. Computer-simulated models depict this structure with accuracy, as confirmed by site-directed mutagenesis. Computer-simulated drug design using computer-simulated models may now be applied to develop new drugs that may enhance the survival of xenografted organs.