Involvement of cyclooxygenase-2 in the potentiation of allyl alcohol-induced liver injury by bacterial lipopolysaccharide.

Bacterial endotoxin (lipopolysaccharide; LPS) augments the hepatotoxicity of a number of xenobiotics including allyl alcohol. The mechanism for this effect is known to involve the inflammatory response elicited by LPS. Upregulation of cyclooxygenase-2 (COX-2) and production of eicosanoids are important aspects of inflammation, therefore studies were undertaken to investigate the role of COX-2 in LPS-induced enhancement of liver injury from allyl alcohol. Rats were pretreated (iv) with a noninjurious dose of LPS or sterile saline vehicle and 2 h later were treated (ip) with a noninjurious dose of allyl alcohol or saline vehicle. COX-2 mRNA was determined by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR), and liver injury was assessed from activities in serum of alanine and aspartate aminotransferases (ALT and AST, respectively) and from histology. Liver injury was observed only in rats cotreated with LPS and allyl alcohol. Serum ALT activity was increased by 4 h after administration of LPS and continued to increase through 8 h. COX-2 mRNA was detectable at low levels in livers from rats receiving only the vehicles at any time up to 8 h. Expression of COX-2 mRNA was increased by 30 min after administration of LPS and remained elevated through 6 h. Allyl alcohol treatment alone caused an increase in COX-2 mRNA at 4 h (2 h after allyl alcohol) that lasted less than 2 h. In livers from rats cotreated with LPS and allyl alcohol, levels of COX-2 mRNA were greater than levels seen with either LPS or allyl alcohol alone. The increased expression of COX-2 mRNA was accompanied by an increase in the concentration of prostaglandin (PG) D(2) in plasma. Plasma PGD(2) concentration was increased to a greater extent in rats treated with LPS plus allyl alcohol compared to allyl alcohol or LPS alone. Pretreatment with the COX-2 selective inhibitor, NS-398, abolished the increase in plasma PGD(2) and reduced the increase in ALT and AST activities observed in rats cotreated with LPS and allyl alcohol. NS-398 did not affect liver injury from allyl alcohol alone administered at a larger, hepatotoxic dose. In addition, ibuprofen, a nonselective inhibitor of cyclooxygenases, did not protect against liver injury from LPS plus allyl alcohol. In isolated hepatocytes PGD(2), but not PGE(2), reduced the concentration of allyl alcohol required to cause half-maximal cytotoxicity. These results suggest that products of COX-2 play a role in the augmentation of allyl alcohol-induced liver injury by LPS.

Physicians' reliance on specialists, therapists, and vendors when prescribing therapies and durable medical equipment for children with special health care needs.

OBJECTIVES: Physicians who care for children with special (health care) needs (CWSN) often must prescribe therapies and/or specialized, durable medical equipment (DME). Given this responsibility and the increasing scrutiny of prescribing practices by various oversight agencies, understanding the extent to which pediatricians rely on their own expertise when prescribing therapies and DME is an important area of research. METHODS: As part of an ongoing investigation of physician preparedness for and practice in prescribing therapies, DME, or procedures for CWSN, we mailed surveys to practicing pediatricians in each of 2 states-Ohio and Mississippi-and to a senior resident at all identified pediatric residency-training programs. The surveys polled recipients as to who they would rely on themselves-specialists, therapists or vendors-to make prescription decisions for a variety of therapies and DME of increasing complexity. We report results as proportions of returned and completed questionnaires. Comparisons among the 3 groups (pediatricians from Ohio and Mississippi and residents) were made with the use of chi(2) analysis. RESULTS: For some categories of therapy and DME, physicians and residents reported that they would take an active role in prescription decisions, and their reliance on specialty consultation increased appropriately with the increasing complexity of the device or therapy. However, respondents generally seemed to share responsibility rather than rely on themselves as sole decision makers for most categories; fewer than one fourth took sole responsibility. Reliance on nonphysician health care providers was evident for all categories; in some cases, up to half of the respondents would allow therapists to take over these decisions, and a small but significant percentage of physicians would entrust DME prescription decisions to vendors alone. CONCLUSIONS: Our findings indicate that many practicing pediatricians and those in training may be unwilling to assume sole responsibility in prescribing and managing therapies and DME for CWSN. Although the number who would rely on consultation with specialists is somewhat reassuring, we found that a significant percentage would turn to nonphysician health care providers and even vendors to make these decisions in some cases, raising liability implications, conflict-of-interest issues, and quality-of-care issues. To protect themselves and their patients from fraud and inappropriate prescriptions and medical management, pediatricians must become increasingly conscientious about complying with American Medical Association guidelines and federal and state laws regarding initiation and supervision of therapies and DME. We offer some recommendations that may help to address this problem.

Protective helmets for children with special health care needs.

An illustrative case of a second traumatic brain injury in the same child raises the issue of the criteria by which protective helmets for disabled children should be prescribed. There is a dearth of data substantiating criteria for helmet prescription for special needs children and assessing potential adverse side effects of routinely wearing a helmet. Further studies seem warranted to determine whether protective helmets truly reduce the frequency and severity of secondary trauma and whether there are adverse effects that outweigh benefits.

Pentoxifylline attenuates bacterial lipopolysaccharide-induced enhancement of allyl alcohol hepatotoxicity.

Small amounts of exogenous lipopolysaccharide (LPS) (10 ng/kg-100 microg/kg) enhance the hepatotoxicity of allyl alcohol in male Sprague-Dawley rats. This augmentation of allyl alcohol hepatotoxicity appears to be linked to Kupffer cell function, but the mechanism of Kupffer cell involvement is unknown. Since Kupffer cells produce tumor necrosis factor-alpha (TNF alpha) upon exposure to LPS, and this cytokine has been implicated in liver injury from large doses of LPS, we tested the hypothesis that TNF alpha contributes to LPS enhancement of allyl alcohol hepatotoxicity. Rats were treated with LPS (10-100 microg/kg iv) 2 h before allyl alcohol (30 mg/kg ip). Co-treatment with LPS and allyl alcohol caused liver injury as assessed by an increase in activity of alanine aminotransferase in plasma. Treatment with LPS caused an increase in plasma TNF alpha concentration, which was prevented by administration of either pentoxifylline (PTX) (100 mg/kg iv) or anti-TNF alpha serum (1 ml/rat iv) one h prior to LPS. Only PTX protected rats from LPS-induced enhancement of allyl alcohol hepatotoxicity; anti-TNF alpha serum had no effect. Exposure of cultured hepatocytes to LPS (1-10 microg/ml) or to TNF alpha (15-150 ng/ml) for 2 h did not increase the cytotoxicity of allyl alcohol (0.01-200 microM). These data suggest that neither LPS nor TNF alpha alone was sufficient to increase the sensitivity of isolated hepatocytes to allyl alcohol. Furthermore, hepatocytes isolated from rats treated 2 h earlier with LPS (i.e., hepatocytes which were exposed in vivo to TNF alpha and other inflammatory mediators) were no more sensitive to allyl alcohol-induced cytotoxicity than hepatocytes from naïve rats. These data suggest that circulating TNF alpha is not involved in the mechanism by which LPS enhances hepatotoxicity of allyl alcohol and that the protective effect of PTX may be due to another of its biological effects.

Training of pediatricians in care of physical disabilities in children with special health needs: results of a two-state survey of practicing pediatricians and national resident training programs.

OBJECTIVE: One goal of the American Academy of Pediatrics' Future of Pediatric Education II Project is to establish guidelines in training physicians to care for children with special health care needs (CWSN). Assessment of current practices in prescribing therapies and devices is necessary to meet this goal. Although much has been written about CWSN, there is a paucity of literature describing pediatricians' preparedness in prescribing such therapies and devices to children with physical disabilities. In an effort to assess physician preparedness, we surveyed pediatric residents nationwide and practicing pediatricians from 2 states, 1 urban and 1 rural. METHODS. A questionnaire aimed at identifying areas of concern regarding preparedness of physicians in practice and in training was prepared and mailed to prospective participants in Ohio and Mississippi. After follow-up mailings to nonresponders, approximately 59% responded. Summary statistics were reported as proportions with 95% confidence intervals. RESULTS: Among those polled, >70% reported no training in prescribing certain durable medical equipment and over 50% reported no training in prescribing certain therapies. In addition, at least 20% reported no training in treating some of the more common childhood physical disabilities. Nearly three fourths of the respondents indicated that they did not believe that they were adequately prepared to take an active role in prescribing therapies and durable medical equipment. Fewer respondents believed that they should be the sole providers of these therapies and durable medical equipment. CONCLUSIONS: The results of the survey indicate a lack of specific training and physician confidence in prescribing therapies and devices to CWSN, establishing the necessity of expanding training programs to better ensure quality health care for special needs children. Although additional ongoing research is necessary to fully evaluate the preparedness of physicians in caring for CWSN, this survey does help to identify areas of physician training that require improvement to provide quality health care for CWSN.

Unrestricted riding in pickup truck cargo beds poses significant hazards.

Individuals riding in the beds of pickup trucks face significant risks of debilitating injury or death, yet Mississippi currently has no legislation restricting ridership in truck beds. Data collection on accidents involving truck bed passengers indicates that children make up the majority of victims. Such accidents impose a heavy burden on society in terms of both medical expenses and impaired quality of life for the victims.

Bacterial endotoxin enhances the hepatotoxicity of allyl alcohol.

Lipopolysaccharide (LPS), or bacterial endotoxin, causes liver damage at relatively large doses in rats. Smaller doses, however, may influence the response to other hepatotoxicants. The purpose these studies was to examine the effect of exposure to relatively all doses of LPS on the hepatotoxic response to allyl alcohol, which causes periportal necrosis in laboratory rodents through an known mechanism. Rats were pretreated with LPS (100 micrograms/kg) 2 hr before treatment with a minimally toxic dose of allyl alcohol mg/kg), and liver toxicity was assessed 18 hr later from activity of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in plasma and from histologic changes in liver sections. Plasma ALT and AST activities were not elevated significantly in rats treated with vehicle, LPS, or allyl alcohol alone, but pronounced increases were observed in rats treated with LPS and allyl alcohol. Significant liver injury occurred as early as 2 hr after allyl alcohol treatment in LPS-pretreated rats and peaked at 6 hr. LPS treatment did not affect the activity of alcohol dehydrogenase and did not affect the rate of production of NADH in isolated livers perfused with allyl alcohol; thus, LPS does not appear to increase the metabolic bioactivation of allyl alcohol into acrolein. On the other hand, pretreatment with 4-methylpyrazole, an inhibitor of alcohol dehydrogenase, abolished the hepatotoxicity of allyl alcohol in LPS-treated rats, indicating that production of acrolein was needed for LPS enhancement of the toxicity of allyl alcohol. Pretreatment of rats with gadolinium chloride (10 mg/kg), a known inactivator of Kupffer cell phagocytic function, decreased LPS augmentation of the response to allyl alcohol. These data indicate that LPS markedly enhances the hepatotoxic response to allyl alcohol. Furthermore, the results suggest that the LPS-induced enhancement of allyl alcohol hepatotoxicity occurs through a Kupffer cell-dependent mechanism.

In vivo cancer diagnosis of the esophagus using differential normalized fluorescence (DNF) indices.

BACKGROUND AND OBJECTIVE: We report the use of new diagnostic parameters based on the differential normalized fluorescence (DNF) signals for malignant tumor diagnosis. STUDY DESIGN/MATERIALS AND METHODS: Over 200 measurements of endogenous fluorescence from normal and malignant esophageal tissues were performed during routine endoscopy in 48 patients. A pulsed nitrogen-pumped dye laser was used to provide in situ excitation at 410 nm. Direct collection of the fluorescence signal emitted by the tissue was achieved using an intensified photodiode array detector equipped with a fiberoptic probe. RESULTS: The fluorescence signals were normalized with respect to the total fluorescence signal area. The cancer diagnosis indices were defined by the difference between the normalized fluorescence signal of a tumor and the mean value of a reference set of normal tissues. The results of the DNF approach were compared with endoscopic examinations and histopathology interpretations of the biopsy samples. Excellent correlation in the classification of normal and malignant tumors for the samples was found. CONCLUSION: The data indicated that the DNF approach has a significant potential to provide a direct, real-time, and in-situ technique for cancer diagnosis of the esophagus without requiring biopsy of the tumors and time-consuming histopathology tests.

Hyperpyrexia associated with sustained muscle contractions: an alternative diagnosis to central fever.

Muscle activity is the principal source of body heat production, and elevated core body temperatures may occur in healthy exercising persons. Hyperpyrexia from sustained tonic muscle contractions can also occur in a number of pathological conditions. The present case of hyperpyrexia associated with dystonic posturing and sustained muscle contraction in a child with encephalopathy illustrates the importance of recognizing muscular activity in the generation of fever of unknown origin following central nervous system injury. The pathophysiology, clinical features, and management of this uncommon cause of fever are discussed.

Natural killer cells induce activated murine B cells to secrete Ig.

We previously demonstrated that dextran-conjugated anti-IgD antibodies (alpha delta-dex) induce proliferation of small, B cell-enriched murine spleen cells (Be cells), and in the presence of IL-2, stimulate Ig secretion in vitro. We have shown that alpha delta-dex-stimulated B cells provide an in vitro model for studying B cell activation by T cell-independent type 2 (TI-2) Ag, as exemplified by the bacterial polysaccharides. We now show that highly purified resting B cells, obtained by electronic cell sorting (Bsp cells), fail to secrete Ig in the presence of alpha delta-dex + IL-2. The alpha delta-dex + IL-2-induced Ig secretory response of Bsp cells is restored upon addition of splenic non-B, non-T cells or a pure population of in vitro-generated NK cells. Similarly, pretreatment of Be cells with anti-AsGm-1 plus complement inhibits Ig secretion in response to alpha delta-dex + IL-2. An IL-2-induced NK cell supernatant (NKSN) is equally potent at stimulating Ig secretion by alpha delta-dex-activated Bsp cells, indicating that cell contact between Bsp and activated NK cells is not required for this effect. IL-2 stimulates not only NK cells, but B cells as well, since addition of anti-IL-2 + anti-IL-2R antibodies to Bsp cell cultures, in the presence of alpha delta-dex + NKSN, inhibits Ig secretion. These data describe a novel animal model for NK cell-induced B cell maturation to Ig secretion and suggest a pathway for Ig production in response to T1-2 Ag.

Comparative in vitro analysis of proliferation, Ig secretion, and Ig class switching by murine marginal zone and follicular B cells.

We have previously demonstrated that activation of murine B cells by dextran-conjugated anti-IgD antibodies may serve as a polyclonal, in vitro model system for studying immune responses to T cell-independent type 2 (TI-2) Ag, as exemplified by the bacterial polysaccharides. Because in vivo Ig responses to TI-2 Ag are mediated primarily by B cells resident in the splenic marginal zone, we wished to determine whether this reflected an intrinsic difference in the responsiveness of marginal zone B cells (MZB) compared with follicular B cells (FB) to this class of Ag. In this report we demonstrate that highly purified MZB, isolated by electronic cell sorting, exhibit a lower proliferative response in vitro in response to unconjugated anti-Ig antibody as well as to dextran- or Sepharose-conjugated anti-IgM or anti-IgD antibodies, whereas they proliferate equal to or better than FB when stimulated by other B cell mitogens including LPS, Salmonella typhimurium mitogen, or anti-CD3-activated CD4+ Th2 cell clone. Despite the different proliferative responses of MZB and FB induced by anti-Ig, Ag receptor cross-linkage stimulates comparable increases in intracellular free calcium concentrations in both of these B cell populations. Furthermore, MZB secrete Ig and undergo Ig isotype switching to a comparable degree, relative to FB, in response to both T cell-dependent and T cell-independent stimuli. This suggests that the compartmentalization of TI-2 responses to the splenic marginal zone rather than the follicular zone reflects something other than the intrinsic responsiveness of the B cells from these two sites.

Comparative study between pulsed and continuous wave lasers for Photofrin photodynamic therapy.

A study was conducted in the normal canine esophagus to compare continuous wave (CW) and pulsed laser light for photodynamic therapy with Photofrin (4 mg/kg). Forty-eight hours post-injection, 630 nm laser light (CW light from an argon-pumped dye-laser and pulsed light from a KTP/532-pumped dye-laser) was delivered using a 24 mm diameter cylindrical esophageal PDT balloon positioned at either distal or proximal esophagus. A 1.0 cm cylindrical diffuser placed in the center of the balloon delivered 300 J/cm of light at an intensity of 400 mW/cm. Three dogs received CW light proximally and pulsed light distally. Four dogs received CW light distally and pulsed light proximally. The light dose delivered to the esophageal mucosa was measured using three isotropic probes placed on the balloon wall. Laser-induced fluorescence technique was used to compare photosensitizer fluorescence intensities at distal and proximal locations. Similar mucosal light doses and drug fluorescence intensities were verified for sites receiving pulsed or CW laser light. Two days after light delivery, the dogs were endoscoped to evaluate the severity of the lesions. While some response variability was observed among different animals, endoscopic examination of the lesions revealed comparable injury from CW and pulsed light in each subject. The animals were then euthanized and necropsies were performed. Based on the gross and histological examination of the lesions, the CW and pulsed laser-induced injuries could not be distinguished.

Quantification of phthalocyanine concentration in rat tissue using laser-induced fluorescence spectroscopy.

Quantification of photosensitizer concentration in tissue should improve planning and outcome of photodynamic therapy. Laser-induced fluorescence (LIF) can be used to measure in vivo fluorescence of photosensitizers in tissue. This study was designed to determine if in vivo fluorescence intensity of chloroaluminum phthalocyanine tetrasulfonate correlates with its concentration in different rat tissues. Following LIF measurements, the animals were humanely euthanized and the concentration of phthalocyanine in different tissues was determined using chemical extraction technique. The correlation of phthalocyanine fluorescence intensity and its concentration was determined for each tissue using Pearson product-moment correlation analysis. A strong correlation between in vivo phthalocyanine fluorescence intensity and its concentration was found for spleen, kidney, liver, and chemically induced mammary adenocarcinoma. Low correlation was found for thigh skin and planum of nose. No correlation was found for thigh muscle and tongue.

Centering balloon to improve esophageal photodynamic therapy.

A cylindrical balloon was developed to improve delivery of circumferential light for photodynamic therapy (PDT) of esophageal carcinoma. The balloon consisted of a 36-mm-long clear cylindrical membrane and a central tube to hold a cylindrical diffuser in the center of the lumen. Three isotropic probes were placed on the outside of the balloon to allow measurement of delivered light dose to the esophageal mucosa. The balloon was tested in the normal esophagus of 8 dogs that were injected with 4.0 mg/kg of PHOTOFRINR. Endoscopy was performed 48 hours following the injection, and under endoscopic observation the balloon assembly was passed, fixed in place, and inflated. A 1-cm cylindrical diffuser was passed into the central tube and 150, 300, and 600 Joules/cm of 630 nm laser light was delivered at 25 cm, 15 cm, and 5 cm proximal to the gastroesophageal junction. One control dog was illuminated using the cylindrical diffuser alone at doses of 300 and 600 Joules/cm of diffuser. Complete circumferential tissue response was obtained when the balloon was used. Relatively uniform light intensities were measured around the lumen. In contrast, noncircumferential and unpredictable PDT responses were generated when the cylindrical diffuser was used without the balloon.

The Alabama automated defibrillation pilot program.

The Alabama Chapter of the American College of Emergency Physicians is now sponsoring a pilot program using EMT-Basic equipped with semi-automatic ("smart") defibrillators for the treatment of cardiac arrest patients in the field. This article outlines the rationale for the program along with present status of results.

Quantitation of the host cell infiltration kinetics of the nonimmunogenic colon 26 tumor by multiparameter flow cytometry.

In this study, we examined the kinetics of host cell infiltration into the nonimmunogenic Colon 26 tumor. We found that 1 x 10(4) cells were required to produce tumors in 100% of mice. The vivo doubling time was 42.5 h, and a barely palpable tumor contained 8 x 10(6) cells. No evidence of concomitant immunity was found. The number of host cells infiltrating the in vivo tumors increased at the same rate as the number of tumor cells, but averaged only 22% of total cells. Cycling T lymphocytes were present in the host cell infiltrate of this tumor. In addition, approximately 50% of in vivo Colon 26 cells were Thy-1.2 positive. The observed characteristic of low immunogenicity makes it a useful murine model for studying human malignant tumors.