Ellen Heck

Comparative study of descemet stripping automated endothelial keratoplasty donor preparation by Moria CBm microkeratome, horizon microkeratome, and Intralase FS60.

Purpose: To report a rare case of large conjunctival B-cell lymphoma in a child. Methods: A 13-year-old girl was initially diagnosed with a right lower eyelid chalazion. After 3 weeks during which the mass was growing, she was referred for treatment to our department. Because of the unusual appearance of the mass, an excisional biopsy was performed. Results: Pathological findings were consistent with those of a large B-cell lymphoma. CD20 and Ki67 staining were positive, and polymerase chain reaction analysis showed monoclonality of B cells. Conclusions: Although conjunctival lymphoma is a very rare entity in children, it should be included in the differential diagnosis of an eyelid or conjunctival mass.PURPOSE To evaluate the quality of stromal bed and the safety on endothelium in preparation of donor tissue for Descemet stripping automated endothelial keratoplasty in a masked fashion using 2 mechanical microkeratomes and a femtosecond laser. METHODS Deep anterior lamellar dissection was performed on 15 donor corneas. Central endothelial cell density was calculated using specular microscopy before and after the dissection. One cornea from each of 5 donor pairs was cut with the Moria ALTK system with the CBm microkeratome using the 300-μm head and the mate cut with the Horizon disposable 300-μm microkeratome. Five additional donor corneas were cut with the Intralase 60-kHz FS laser. The donor corneas were then bisected with half of the cornea used for Live/Dead assay to study central endothelial viability. The other halves were sent for scanning electron microscopy of the stromal bed. Qualitative surface roughness of the scanning electron microscopy images was graded by 2 masked observers, and quantitative surface roughness was assessed using roughness evaluation software. RESULTS The Horizon group showed a smoother stromal bed compared with the Moria or Intralase groups by 2 masked observers. However, the Moria group had the smoothest quantitative score of all the groups when assessed by roughness evaluation software. There was no statistically significant difference among the 3 groups in the percentage change in the central endothelial cell density or percentage of viable central endothelium by Live/Dead assay after the dissection. CONCLUSIONS Both mechanical microkeratomes created smoother stromal bed dissections than the femtosecond laser. All systems provided good endothelial cell viability.

A critical evaluation of povidone-iodine absorption in thermally injured patients.

Povidone-iodine ointment is a widely used topical antimicrobial agent in thermally injured patients. In 17 patients with burns ranging from 4 to 85% TBSA this agent was applied to both partial- and full-thickness burns. Peak serum iodine levels in patients treated within 24 hours of injury ranged from 595 to 4,900 micrograms per dL. The amount of iodine absorbed was directly related to the size of the burn. Serum iodine levels continued to rise until the drug was discontinued and remained elevated for as long as 7 days after discontinuance. Iodine excretion was directly related to renal function. The highest serum and lowest urinary iodine levels were present in patients who developed renal failure. Thyroid function was not affected by drug usage. A proven cause-and-effect relationship between the elevated serum iodine attributed to the absorption of povidone-iodine and metabolic acidosis, hypernatremia, and hyperosmolarity was not established. Close monitoring of the patients fluid and electrolyte balance is imperative during drug usage. The drug is contraindicated in any patient with impaired renal function. The high serum levels of this inorganic ion imply potential toxicity, but clinical evidence of cell or organ toxicity is as yet undetermined.

A comparison of leukocyte function and burn mortality.

Of the several tests which measure leukocyte bactericidal competency only one, chemotaxis, has been reported to reflect both leukocyte dysfunction and patient mortality in major thermal injury. To validate the reliability that chemotaxis reflects mortality functional chemotactic index (FCI) was measured in leukocytes from 33 patients with 30% or greater total body surface area burns. Additionally, nitroblue tetrazolium (NBT) reduction and O2 consumption were compared to FCI. Significant decreases were seen in FCI values compared to normals. No differences, however, were seen in FCI values between surviving and nonsurviving patients. NBT reduction and O2 consumption also showed decreases with a significant difference between the mean decrease in surviving and nonsurviving patients. While the data show all these laboratory tests to be reflective of significant impairments in host defense, they do not appear to be reliable as predictive indices of patient survival and therefore should not be interpreted as such.

The role of NADH-NADPH oxidase activity in the leukocyte function of burned patients

Irreversible sepsis, in spite of advancements in topical therapy and antimicrobial agents, remains the leading cause of death in major thermal injury. A defect in intracellular bactericidal capacity in leukocytes from severely burned patients appears to correspond with increases in bacterial wound colonization and ultimate sepsis. This leukocyte defect has been demonstrated by abnormally low nitroblue tetrazolium reduction (NBT) and oxygen consumption of white cells in patients with major thermal injury. The subcellular mechanisms responsible for decreased bactericidal capacity were therefore investigated. Nicotinamide-adenine dinucleotide (NADH) and nicotinamide-adenine phosphodinucleotide (NADPH) oxidase activity was measured in patients with major burns, controls (normals), and in patients with nonburn stress or infection. NADH and NADPH oxidase levels in leukocytes from burn patients were not significantly different from those of normal nonchallenged controls but were significantly lower than the leukocyte values found in the patients with nonburn infections or stress. This NADH and NADPH defect in the subcellular leukocyte fraction suggests that it may be a significant factor in the reduced bactericidal function of the intact leukocyte in thermally injured patients.

Usefulness of blood cultures in confirming septic complications in burn patients: evaluation of a new culture method.

A prospective evaluation of several techniques used to monitor thermally injured patients for septic complications was initiated. Thirty-eight extensively burned patients, at high risk for septic complications, were selected for study. Of these, 92% developed significant burn wound infection. Careful daily clinical evaluation and serial quantitative burn wound biopsy cultures provided the most effective means of establishing an early diagnosis of would sepsis. Serial blood cultures performed by the two different techniques were of very limited value in the recognition were not recovered until 5-10 days following documentation of heavy bacterial colonization of the burn wound.

ELISA HIV testing and viral culture in the screening of corneal tissue for transplant from medical examiner cases

The spread of AIDS and its association with a retrovirus, commonly called T-lymphotropic virus type III (HIV) has produced great concern among those involved in transplantation about the possibility for transmission of this disease through tissue and organ transplant. Isolation of the virus from conjunctiva and tears further heightened this concern for corneal transplants. This review of medical examiner (ME) cases, a population in which high numbers of transplant corneas are retrieved, has revealed positive serologic tests for HIV in serum from 5 cases in 205 screens. These represent potential donors based on other standard medical criteria available at the time of collection. This tissue was also submitted for viral culture, and virus was recovered from the corneal tissue in McCarey-Kaufman (MK) media. These findings and case reviews are discussed.

Viral HCV RNA reactivity of corneal cells in plasma HCV nucleic acid-positive eye donors.

Purpose: Hepatitis C virus (HCV) infection is a worldwide health concern according to the World Health Organization, and a positive serology test for HCV is currently one of the most common reasons for tissue rejection for transplantation. Demonstrating the presence or absence of viral RNA or DNA in corneal cells, a privileged avascular site, verifies the importance of screening and testing protocols, which eliminate individual donor tissue from potential release for transplantation. Methods: Improved tests such as nucleic acid test (NAT) have added a significant advantage in the arsenal of measures employed to determine donor eligibility. Standard testing for blood using nucleic acid technology was applied to aliquots of corneal cells obtained by scraping pooled epithelial and endothelial cells from individual donor corneas. Results: Results of the current study further confirm and extend the importance of NAT blood tests by the demonstration of a high percentage (77%) of corneal cells that test positive with HCV NAT in conjunction with concomitant serum-positive ocular tissue donor samples. Conclusions: These data provide important support for Center for Disease Control and Prevention reports that state that 3 of 4 seropositive individuals were also viremic, and reemphasize the importance of routine NAT blood testing in assuring safe tissue transplants.

Guidelines for preventing 'dilution false negatives' in in vitro laboratory testing of the donor population

In this article we develop a calculation or formula for use in determining the potential dilution effect of fluids administered during patient treatment on serologic testing parameters. The formula uses basic principles of (a) fluid distribution over time from administration; (b) ratios of plasma and extravascular fluid volumes to body weight; and (c) common practices of fluid resuscitation. A dilution threshold of 50% was set using data from enzymelinked immunosorbent assay human immunodeficiency virus antibody determinations performed on in vitro diluted seropositive serum samples. These data respond to issues raised by guidelines from the Centers for Disease Control and the U.S. Food and Drug Administration to achieve recipient safety without unnecessarily restricting the potential donor pool.

Effect of Circulating Fibronectin on Stimulation of Leukocyte Oxygen Consumption and Serum Opsonizing Function in Burned Patients

In a study of 27 thermally burned patients (mean TBSA, 58%; range, 32-96%) serum fibronectin levels were decreased with parallel decreased oxygen consumption of stimulated peripheral blood phagocytes and decreased EGTA-blocked burn serum opsonizing activity which correlated with serum fibronectin changes postburn. Normal and burn sera fibronectin content also correlated with the opsonizing times for zymosan and Staphylococcus aureus but not for Enterobacteriaceae. Although in vivo 14 cases showed circulating fibronectin 140 micrograms/ml or lower and a marked decrease in Staphylococcus aureus opsonization, only two patients from this group revealed positive Staphylococcus aureus blood cultures and serum fibronectin levels were higher in patients with Staphylococcus aureus sepsis than in patients with Enterobacteriaceae sepsis. Supplementary experiments on leukocyte oxidative response after zymosan stimulation in normal, fibronectin-depleted, and fibronectin-reconstituted serum demonstrated that the lag period of oxygen burst is a fibronectin-dependent reaction.

The Importance of the Bacterial Flora in Cadaver Homograft Donor Skin: Bacterial Flora in Cadaver Homograft

The increased use of cadaveric homograft skin, made possible through the development of skin banking techniques, has emphasized the necessity of bacteriologic control of transplant tissue. Since the inoculation of burn wounds with pathogenic organisms is known to be potentially lethal, the lack of adequate quality assurance programs for controlling donor graft flora could represent a significant hazard. This potential hazard was evaluated by studying bacterial flora isolates from 362 continuous cadaver donors harvested during an 18-month period. Positive cultures were found in 169 (46.6%) donor skins. Fifty-one (14%) of the samples grew a variety of potentially pathogenic organisms. Included in the potential pathogenic isolates were 36 aerobic gram-negative rods, three gram-positive cocci, and eight anaerobic organisms, six Clostridial species, and two Bacteroides species. All 51 of these donations were rejected for transplant. The remaining 118 cases grew indigenous skin flora and were accepted for transplant. These results emphasize the importance of routine culture assessment of all donor homografts before transplant. Additionally, the unexpected finding of anaerobic organisms (unpredicted by preculture donor screening) in the donor population indicates a need for a system of isolation and identification of uncommon or unexpected skin flora.