Whole-Blood Assay to Measure Oxidative Burst and Degranulation of Neutrophils for Monitoring Trauma Patients

AbstractBackground and Purpose: Polymorphonuclear neutrophils (PMNs) protect the host from invading microorganisms, but excessive PMN activation after trauma causes tissue injury. Rapid monitoring of PMN function is critical for the assessment of the inflammatory state of trauma patients. Here, the authors adapted two simple and rapid methods to measure oxidative burst and degranulation of human PMNs in whole blood to avoid potential interference of cell isolation procedures with the assessment of PMN function.Material and Methods: Heparinized blood was drawn from healthy volunteers or trauma patients, preincubated at 37 °C for 5 min, and stimulated with N-formyl-methionyl-leucyl-phenylalanine (fMLP). Four assays for oxidative burst were tested: (1) cytochrome C; (2) homovanillic acid (HVA); (3) Amplex^® Red; and (4) flow cytometry with dihydrorhodamine 123 (DHR). PMN degranulation was assessed with flow cytometry using antibodies to: (1) CD11b/Mac-1 (CD18); (2) CD63; and (3) CD66b (CD67).Results: With the exception of the DHR method, all methods to measure oxidative burst were found to be unsuitable in whole blood due to interference of plasma proteins and hemoglobin with the fluorimetric or photometric readouts. By contrast, all degranulation methods were suitable for whole-blood studies. However, for the assessment of formyl peptide-induced degranulation, anti-antibodies to CD11b/Mac-1 and CD66b were up to five times more sensitive than antibodies to CD63. Thus, the degranulation and DHR methods were optimized for increased sensitivity, speed, and specificity and their usefulness to measure PMN function in trauma patients was tested.Conclusion: The whole-blood methods based on flow cytometry with DHR, anti-CD11b/Mac-1, and anti- CD66b are rapid, simple, and reliable techniques to assess PMN function for trauma research.     

Effect of Mutans streptococcal colonization on plaque formation and regrowth in young children--a brief commnunication.

OBJECTIVES: Visible plaque on the maxillary anterior teeth of young children has been identified as a risk indicator for early childhood caries. The present study examined whether this plaque is related to the colonization of children's teeth with Mutans streptococci (MS) or toothbrushing frequency. METHODS: Thirty-nine children, aged 12 to 36 months, had plaque scores, and plaque samples were taken from the labial surfaces of the maxillary incisors at baseline and repeated 3 days after suspended oral hygiene (plaque regrowth). RESULTS: A positive correlation was found between the baseline percent MS and regrowth plaque score and between baseline and regrowth plaque scores. Plaque scores of those that brushed zero to one time a day were not different from those who brushed two or more times a day. CONCLUSIONS: This study suggests that the presence of plaque on the anterior teeth of young children is consistent and related to MS colonization.     

Retinal degeneration associated with RDH12 mutations results from decreased 11-cis retinal synthesis due to disruption of the visual cycle

Human Molecular Genetics     

Effects of ketotifen 0.025% and lodoxamide 0.1% on eosinophil infiltration into the guinea pig conjunctiva in a model of allergic conjunctivitis.

The effects of ketotifen and lodoxamide on eosinophil infiltration were assessed in a guinea pig model of allergic conjunctivitis. The two active treatments were coded in this masked study in which 30 male guinea pigs, sensitized to chicken egg albumin (ovalbumin), were randomly assigned to one of three groups: Group 1, instillation of 0.9% NaCl into the conjunctival sac of both eyes; Group 2, instillation of 0.025% ketotifen into the left eye and 0.9% NaCl into the right eye; Group 3, instillation of 0.1% lodoxamide into the left eye and 0.9% NaCl into the right eye. Ovalbumin was administered topically to each eye, except in Group 1 where it was only applied to the left eye. (111)In-oxine labeled eosinophils were injected into the jugular vein of each guinea pig; the animals were sacrificed 17 hours after ovalbumin had been applied. The level of radioactivity in the ketotifen- and lodoxamide-treated eyes was approximately 60% of that in the saline-treated eyes. Moreover, the mean level of radioactivity in the ketotifen- and lodoxamide-treated eyes was comparable with the mean level of radioactivity in the saline-treated eye of Group 1, which had not been exposed to allergen. These results indicate that the therapeutic effects of ketotifen and lodoxamide in allergic conjunctivitis may be partly mediated by an inhibitory effect on eosinophils.     

Ethylene oxide does not extinguish the osteoinductive capacity of demineralized bone: A reappraisal in rats

We examined the influence of ethylene oxide (EO) and gamma irradiation on the osteoinductive capacity of demineralized bone. Demineralized bone powder prepared from Wistar rats was exposed to EO (55 °C or 40 °C) or gamma irradiation (25 KGy) or was preserved in ethanol. Sterilely-prepared bones served as controls. The powder was packed in a gelatin capsule and implanted for 6 weeks in muscles of 6-week-old female rats. Exposure of demineralized bone particles to EO 55 °C resulted in an almost complete loss of osteoinductivity. Irradiated bones lost about 40% of their osteoinductive capacity, while sterilization with EO at 40 °C resulted in only a slight alteration of the osteoinductivity, as assessed by the recovered weight ratio, calcium content, alkaline phosphatase activity measurements and histo-morphometry. Ethanol treatment had no influence on the new bone yield when compared to controls.

As EO exposure at 40 °C is a true sterilization procedure, it can be recommended in a clinical setting for its small effect on osteoinductive capacity as assessed experimentally in rats.     

Narrative and procedural discourse in temporal lobe epilepsy.

It is well established that some individuals with temporal lobe epilepsy (TLE) demonstrate language deficits at the single word level. However, discourse production rarely has been examined quantitatively within this group. This study compared adult TLE patients with an early seizure onset (< or = age 14 years, n = 27) to a control group (n = 28) on narrative and procedural discourse tasks. As a group, the TLE patients performed normally on the procedural discourse task, but differed significantly from the controls on several narrative discourse variables. At the individual level, 30% of the TLE patients versus 4% of the controls demonstrated impaired discourse ability (p and 0.01). Within this early onset TLE group, discourse performance was not associated with demographic or seizure history variables. Considering the cognitive domain, discourse performance correlated significantly with working memory. In summary, mild discourse dysfunction was present in a significant minority of early onset TLE patients, but this deficit was not closely associated with other language measures. Discourse ability and its neuropsychological, neuroanatomical and conversational speech correlates deserve further study in TLE patients.     

Thoracoscopic wedge resection

Summary Thoracoscopic surgery is decidedly expanded by the ability to perform pulmonary wedge resections of the lung by using the Endo-GIA-stapler. In addition to thoracoscopic biopsies, since July 1991 we have carried out wedge resections in 12 patients suffering from spontaneous pneumothorax (nine) or peripheral bronchial carcinoma (three). Postoperatively one air fistula persisted over 9 days. The chest tube was removed within 48 h in all other patients. There was no other major complication. The postoperative hospitalization period lasted 4.6 days (1–9 days). Operating time was 44 min (30–70 min). The benefit for the patient consists in the little-impaired breathing mechanics, the short hospital stay, and the favorable cosmetic result.     

Bromine-76 and carbon-11 labelled NNC 13-8199, metabolically stable benzodiazepine receptor agonists as radioligands for positron emission tomography (PET)

NNC 13-8241 has recently been labelled with iodine-123 and developed as a metabolically stable benzodiazepine receptor ligand for single-photon emission computed tomography (SPECT) in monkeys and man. NNC 13-8199 is a bromo-analogue of NNC 13-8241. This partial agonist binds selectively and with subnanomolar affinity to the benzodiazepine receptors. We prepared ⁷⁶Br labelled NNC 13-8199 from the trimethyltin precursor by the chloramine-T method. Carbon-11 labelled NNC 13-8199 was synthesised by N-alkylation of the nitrogen of the amide group with [¹¹C]methyl iodide. Positron emission tomography (PET) examination with the two radioligands in monkeys demonstrated a high uptake of radioactivity in the occipital, temporal and frontal cortex. In the study with [⁷⁶Br]NNC 13-8199, the monkey brain uptake continued to increase until the time of displacement with flumazenil at 215 min after injection. For both radioligands the radioactivity in the cortical brain regions was markedly reduced after displacement with flumazenil. More than 98% of the radioactivity in monkey plasma represented unchanged radioligand 40 min after injection. The low degree of metabolism indicates that NNC 13-8199 is metabolically much more stable than hitherto developed PET radioligands for imaging of benzodiazepine receptors in the primate brain. [⁷⁶Br]NNC 13-8199 has potential as a radioligand in human PET studies using models where a slow metabolism is an advantage. Received 19 April and in revised form 10 June 1997