Molecular targets of the nitrogen containing bisphosphonates: the molecular pharmacology of prenyl synthase inhibition.

The nitrogen containing bisphosphonates (N-BP) are the drug of choice for treating disease characterised by resorption of bone such as osteoporosis and metastatic bone disease. The overall mechanism of action is achieved through a combination of precise targeting to the bone environment and an extremely potent inhibition of a vital enzyme in an essential metabolic pathway. This targeting to bone is achieved through the phosphate-carbon-phosphate backbone of the drug which gives a high affinity for bone mineral. Once bound to bone the N-BP can be internalised by osteoclasts as they resorb bone where the drug can then interact with its molecular target. The enzyme target of these drugs, FPP synthase, is at a branch point in the mevalonate pathway. This pathway is principally used for the manufacture of cholesterol but also many other biochemicals including farnesyl pyrophosphate and geranylgeranyl pyrophosphate. These prenyl groups are used in the post-transcriptional modification of proteins such as small GTPases that require a lipid membrane anchor to function. The main cellular effect of the blockade of FPP synthase by N-BP is to prevent protein prenylation resulting in disruption to vital signalling pathways and loss of osteoclast function. This review will examine the biochemistry of FPP synthase, inhibition by the N-BP and and other potential uses of prenyl synthase inhibitors.

Differences between bisphosphonates in binding affinities for hydroxyapatite.

Bisphosphonates (BPs) inhibit bone resorption and are widely used for the treatment of bone diseases, including osteoporosis. BPs are also being studied for their effects on hydroxyapatite (HAP)-containing biomaterials. There is a growing appreciation that there are hitherto unexpected differences among BPs in their mineral binding affinities that affect their pharmacological and biological properties. To study these differences, we have developed a method based on fast performance liquid chromatography using columns of HAP to which BPs and other phosphate-containing compounds can adsorb and be eluted by using phosphate buffer gradients at pH 6.8. The individual compounds emerge as discrete and reproducible peaks for a range of compounds with different affinities. For example, the peak retention times (min; mean +/- SEM) were 22.0 +/- 0.3 for zoledronate, 16.16 +/- 0.44 for risedronate, and 9.0 +/- 0.28 for its phosphonocarboxylate analog, NE10790. These results suggest that there are substantial differences among BPs in their binding to HAP. These differences may be exploited in the development of biomaterials and may also partly explain the extent of their relative skeletal retention and persistence of biological effects observed in both animal and clinical studies.

Samplers for evaluation and quantification of ultra-low volume space sprays.

A field study was conducted to explore the suitability of 5 pesticide deposition samplers for airborne spray and ground deposits from ultra-low-volume (ULV) space sprays. Samplers included horizontally stretched stationary cotton ribbons at 2 heights, rotating ribbon, rotating Teflon slides, and filter paper. Slides were also used for droplet-size analysis. A set of 7 samplers of each type was placed at 1, 7, 15, 25, 40, 65, and 90 m from the spray line along the spray swath. Water and BVA13 oil with fluorescent dyes as tracers were sprayed with the use of a truck-mounted ULV sprayer at dusk and dawn. Results suggest that the horizontal and rotating cotton ribbons are best for quantification of airborne spray and filter paper is best for ground deposition collection. The rotating slide samplers only detected the BVA13 oil-based sprays.

Reflectionless tunneling at the interface between nanoparticles and superconductors.

Interfaces between disordered normal (DN) materials and superconductors (S) are known to generate conductance peaks at zero-bias voltage (V) and magnetic field (B). Using molecularly linked Au nanoparticle films as the DN component, we find that superimposed on conductance peaks are oscillations that depend simultaneously on both V and B. Such correlated conductance oscillations are predicted by a "reflectionless tunneling" phenomenon but have not been observed in other DN-S systems. Length scales extracted from periods of conductance oscillation correlate well with film nanostructure.

Identification of a novel phosphonocarboxylate inhibitor of Rab geranylgeranyl transferase that specifically prevents Rab prenylation in osteoclasts and macrophages.

Nitrogen-containing bisphosphonate drugs inhibit bone resorption by inhibiting FPP synthase and thereby preventing the synthesis of isoprenoid lipids required for protein prenylation in bone-resorbing osteoclasts. NE10790 is a phosphonocarboxylate analogue of the potent bisphosphonate risedronate and is a weak anti-resorptive agent. Although NE10790 was a poor inhibitor of FPP synthase, it did inhibit prenylation in J774 macrophages and osteoclasts, but only of proteins of molecular mass approximately 22-26 kDa, the prenylation of which was not affected by peptidomimetic inhibitors of either farnesyl transferase (FTI-277) or geranylgeranyl transferase I (GGTI-298). These 22-26-kDa proteins were shown to be geranylgeranylated by labelling J774 cells with [(3)H]geranylgeraniol. Furthermore, NE10790 inhibited incorporation of [(14)C]mevalonic acid into Rab6, but not into H-Ras or Rap1, proteins that are modified by FTase and GGTase I, respectively. These data demonstrate that NE10790 selectively prevents Rab prenylation in intact cells. In accord, NE10790 inhibited the activity of recombinant Rab GGTase in vitro, but did not affect the activity of recombinant FTase or GGTase I. NE10790 therefore appears to be the first specific inhibitor of Rab GGTase to be identified. In contrast to risedronate, NE10790 inhibited bone resorption in vitro without markedly affecting osteoclast number or the F-actin "ring" structure in polarized osteoclasts. However, NE10790 did alter osteoclast morphology, causing the formation of large intracellular vacuoles and protrusion of the basolateral membrane into large, "domed" structures that lacked microvilli. The anti-resorptive activity of NE10790 is thus likely due to disruption of Rab-dependent intracellular membrane trafficking in osteoclasts.

Outcome study of minors after parental refusal of paramedic transport.

OBJECTIVE: Patient refusal of paramedic transport against medical advice (AMA) has significant medical-legal implications. Previous studies have investigated patient outcomes after refusal of transport, but none has focused on these events in minors. This study was performed to evaluate the outcomes of this patient population after refusal of transport as well as the significance of base hospital physician discussion with parents in the decision to refuse transport. METHODS: This was a retrospective telephone follow-up survey involving parents of minors for whom transport was refused after accessing emergency medical services (EMS) via the 911 system. Data were initially obtained from paramedic run records and each family was subsequently contacted by telephone and surveyed with regard to their experiences with the field medics in addition to the medical follow-up sought for their child and patient outcomes. RESULTS: Eighty-nine patients met criteria for survey. Telephone contact was made with 44 parents, of whom 32 (73% of those contacted, 36% overall) participated. Twenty-seven (84%) received medical follow-up, either at an emergency department or in a private physician's office. Most patients (89%) who were evaluated and/or treated by a physician were subsequently released, while three children were admitted to the hospital, all three with respiratory or cardiac chief complaints. CONCLUSIONS: Children whose parents refused EMS transport received medical follow-up in the majority of cases, with a small group requiring admission.

Exercise-associated hyponatremia in marathon runners: a two-year experience.

This study was conducted to better define the pathophysiology, risk factors, and therapeutic approach to exercise-associated hyponatremia. Medical records from all participants in the 1998 Suzuki Rock 'N' Roll Marathon who presented to 14 Emergency Departments (EDs) were retrospectively reviewed to identify risk factors for the development of hyponatremia. Hyponatremic patients were compared to other runners with regard to race time and to other marathon participants seen in the ED with regard to gender, clinical signs of dehydration, and use of nonsteroidal anti-inflammatory drugs (NSAIDs). An original treatment algorithm incorporating the early use of hypertonic saline (HTS) was evaluated prospectively in our own ED for participants in the 1999 marathon to evaluate improvements in sodium correction rate and incidence of complications. A total of 26 patients from the 1998 and 1999 marathons were hyponatremic [serum sodium (SNa) < or =135 mEq/L] including 15 with severe hyponatremia (SNa < or = 125 mEq/L). Three developed seizures and required intubation and admission to an intensive care unit. Hyponatremic patients were more likely to be female, use NSAIDS, and have slower finishing times. Hyponatremic runners reported drinking "as much as possible" during and after the race and were less likely to have clinical signs of dehydration. An inverse relationship between initial SNa and time of presentation was observed, with late presentation predicting lower SNa values. The use of HTS in selected 1999 patients resulted in faster SNa correction times and fewer complications than observed for 1998 patients. It is concluded that the development of exercise-associated hyponatremia is associated with excessive fluid consumption during and after extreme athletic events. Additional risk factors include female gender, slower race times, and NSAID use. The use of HTS in selected patients seems to be safe and efficacious.

Resuscitation in the out-of-hospital setting: medical futility criteria for on-scene pronouncement of death.

The complete and irreversible cessation of life is often difficult to determine with complete confidence in the dynamic environment of out-of-hospital emergency care. As a result, resuscitation efforts often are initiated and maintained by emergency medical services (EMS) providers in many hopeless situations. Medical guidelines are reviewed here to aid EMS organizations with respect to decisions about: 1) initiating or waiving resuscitation efforts; 2) the appropriate duration of resuscitation efforts; and 3) recommended procedures for on-scene or prehospital pronouncement of death (termination of resuscitation). In cases of nontraumatic cardiac arrest, few unassailable criteria, other than certain physical signs of irreversible tissue deterioration, exist for determining medical futility at the initial encounter with the patient. Thus, the general medical recommendation is to attempt to resuscitate all patients, adult or child, in the absence of rigor mortis or dependent lividity. Conversely, wellfounded guidelines now are available for decisions regarding termination of resuscitation in such patients once they have received a trial of advanced cardiac life support. In practice, however, the final decision to proceed with on-scene pronouncement of death for these patients may be determined more by family and provider comfort levels and the specific on-scene environment. For patients with posttraumatic circulatory arrest, the type of injury (blunt or penetrating), the presence of vital signs, and the electrocardiographic findings are used to determine the futility of initiating or continuing resuscitation efforts. In general, patients who are asystolic on-scene are candidates for on-scene pronouncement, regardless of mechanism. With a few exceptions, blunt trauma patients with a clearly associated mechanism of lethal injury are generally candidates for immediate cessation of efforts once they lose their pulses and respirations. Regardless of the medical futility criteria, specialized training of EMS providers and targeted related testing of operational issues need to precede field implementation of on-scene pronouncement policies. Such policies also must be modified and adapted for local issues and resources. In addition, although the current determinations of medical futility, as delineated here, are important to establish for societal needs, the individual patient's right to live must be kept in mind always as new medical advances are developed.

Optimal prehospital cardiovascular care.

Optimal prehospital cardiovascular care may improve the morbidity and mortality associated with acute myocardial infarctions (AMIs) that begin in the community. Reducing the time delays from AMI symptom onset to intervention begins with maximizing effective patient education to reduce patient delay in recognizing symptoms and seeking assistance. Transportation delays can be minimized by appropriate use of 911 systems and improving technological 911 support. Patient triage to heart centers from the prehospital setting requires strict and comprehensive definition of the criteria for these centers by competent, unbiased clinical societies or governmental agencies. Prehospital 12-lead electrocardiograms and initiation of thrombolytic therapy can provide acute diagnosis and early treatment, thus facilitating faster processing and more directed in-hospital intervention. They also minimize over- and undertriage of patients to cardiac centers. Although evidence from investigational trials suggests that many of these procedures are effective, more research is required to ensure correct implementation and quality assurance at all emergency service levels.

Structure-activity relationships for inhibition of farnesyl diphosphate synthase in vitro and inhibition of bone resorption in vivo by nitrogen-containing bisphosphonates.

It has long been known that small changes to the structure of the R(2) side chain of nitrogen-containing bisphosphonates can dramatically affect their potency for inhibiting bone resorption in vitro and in vivo, although the reason for these differences in antiresorptive potency have not been explained at the level of a pharmacological target. Recently, several nitrogen-containing bisphosphonates were found to inhibit osteoclast-mediated bone resorption in vitro by inhibiting farnesyl diphosphate synthase, thereby preventing protein prenylation in osteoclasts. In this study, we examined the potency of a wider range of nitrogen-containing bisphosphonates, including the highly potent, heterocycle-containing zoledronic acid and minodronate (YM-529). We found a clear correlation between the ability to inhibit farnesyl diphosphate synthase in vitro, to inhibit protein prenylation in cell-free extracts and in purified osteoclasts in vitro, and to inhibit bone resorption in vivo. The activity of recombinant human farnesyl diphosphate synthase was inhibited at concentrations > or = 1 nM zoledronic acid or minodronate, the order of potency (zoledronic acid approximately equal to minodronate > risedronate > ibandronate > incadronate > alendronate > pamidronate) closely matching the order of antiresorptive potency. Furthermore, minor changes to the structure of the R(2) side chain of heterocycle-containing bisphosphonates, giving rise to less potent inhibitors of bone resorption in vivo, also caused a reduction in potency up to approximately 300-fold for inhibition of farnesyl diphosphate synthase in vitro. These data indicate that farnesyl diphosphate synthase is the major pharmacological target of these drugs in vivo, and that small changes to the structure of the R(2) side chain alter antiresorptive potency by affecting the ability to inhibit farnesyl diphosphate synthase.

Are heroin overdose deaths related to patient release after prehospital treatment with naloxone?

OBJECTIVE: Naloxone is frequently used by prehospital care providers to treat suspected heroin and opioid overdoses. The authors' EMS system has operated a policy of allowing these patients, once successfully treated, to sign out against medical advice (AMA) in the field. This study was performed to evaluate the safety of this practice. METHODS: The authors retrospectively reviewed all 1996 San Diego County Medical Examiner's (ME's) cases in which opioid overdoses contributed to the cause of death. The records of all patients who were found dead in public or private residences or died in emergency departments of reasons other than natural causes or progression of disease, are forwarded to the ME office. ME cases associated with opiate use as a cause of death were cross-compared with all patients who received naloxone by field paramedics and then refused transport. The charts were reviewed by dates, times, age, sex, location, and, when available, ethnicity. RESULTS: There were 117 ME cases of opiate overdose deaths and 317 prehospital patients who received naloxone and refused further treatment. When compared by age, time, date, sex, location, and ethnicity, there was no case in which a patient was treated by paramedics with naloxone within 12 hours of being found dead of an opiate overdose. CONCLUSIONS: Giving naloxone to heroin overdoses in the field and then allowing the patients to sign out AMA resulted in no death in the one-year period studied. This study did not evaluate for return visits by paramedics nor whether patients were later taken to hospitals by private vehicles.

Fatal ascending tonic-clonic seizure syndrome.

A fatal case of ascending tonic-clonic seizure (ATCS) syndrome resulted from the inadvertent, unrecognized use of a hyperosmolar ionic contrast agent during myelography. The patient presented with lower-extremity myoclonic jerking, agitation, hyperthermia, rhabdomyolysis, and disseminated intravascular coagulation. Emergency physicians must be cognizant of this unique toxidrome to initiate early, aggressive care.

Differential display analysis of gene expression indicates that age-related changes are restricted to a small cohort of genes.

It is clear that there is a genetic component associated with the ageing process. Although evolutionary theory has suggested that the activity of certain genes may facilitate ageing by favouring resource utilisation by the germ cells at the expense of somatic cells, there is reason to believe that the senescent phenotype, which is the endpoint of the ageing process, may be due to alterations in the levels of expression of other genes. To investigate this situation we have used the differential display technique to survey gene expression during ageing of the rat brain, heart and liver. By optimising this technique it is possible to identify up to 10000-14000 PCR products, which represent genes expressed in the tissue under study. Interestingly, only a relatively small cohort (approximately 2%) of these genes appear to show significant changes in their levels of expression during ageing. Characterisation of the latter has so far revealed certain genes, such as glial fibrillary acidic protein, which are associated with the senescent phenotype. It has also revealed that the level of fos, a component of the AP-1 transcription factor, decreases with age, which has implications for AP-1 regulated genes. The differential display technique has also revealed an increase in mitochondrial RNA during ageing of the heart, which may be due to a gene dosage effect caused by the presence of increased numbers of mitochondrial genomes in myocytes in old age. The differential display technique therefore appears to offer a powerful tool for identifying genes which contribute to the emergence of a senescent phenotype.

Outcome study of prehospital patients signed out against medical advice by field paramedics.

STUDY OBJECTIVE: To describe the incidence and demographic data of prehospital patients who contact paramedics by way of the 911 system, refuse transport against medical advice (AMA), then call 911 and are subsequently reevaluated by paramedics in the following 48 hours. METHODS: We conducted a retrospective observational review of records using the San Diego County Quality Assurance Network database for prehospital providers. All paramedic 911 responses that made base hospital contact over a 3-month period were reviewed to identify patients who signed out AMA. The main outcome measure was to identify patients who signed out AMA and then called 911 again within 48 hours. The demographics, complaints, treatments, and dispositions of these patients are described. RESULTS: Of 6,512 total 911 responses reviewed, 443 (7%) involved patients who signed out AMA. Of these patients, 156 cases (35.2%) were listed as trauma and 287 (64.8%) were medical, with cardiac chest pain, seizure, and respiratory distress/shortness of breath the most frequently noted medical subcategories. Fifty-one (11.5%) such patients received treatment; 34 received dextrose, 12 naloxone, 4 albuterol, and 1 a splint. Patient names were available in 5,515, of the total 6,512 responses and 431 of the 443 AMA cases, permitting computer searching of reevaluations by paramedics. Of the 431 AMA patients for whom a name was available, 10 (2%) called 911 again within 48 hours. All 10 callbacks were made for a related chief compliant, and all 10 of these patients were transported (4 admitted to hospital, 1 died en route, 1 transferred to another facility, 4 discharged from the ED). Of these 10 patients, 7 (70%) were older than 65 years, compared with 17% of all AMA patients older than 65 years. CONCLUSION: On the basis of our findings, patients over the age of 65 years have a propensity to recontact paramedics and should be aggressively encouraged to seek emergency medical treatment. Future prospective studies should be mounted to examine at patient outcome and to assess why patients sign out AMA after making contact with paramedics.

MAST 96.

The Military Anti-Shock Trouser, or MAST suit, is a controversial device that has been used to support blood pressure in hypotensive trauma patients. Most studies on humans have shown that the device has limited clinical utility. In this study, a telephone survey of all 50 State Emergency Medical Services was conducted to determine the nature and extent of MAST suit usage in the United States. The trend in MAST suit usage in San Diego County over the last 7 years was also analyzed. Thirty (60%) states still require MAST suits to be carried on ambulances. In San Diego County, MAST suit inflations for adult, hypotensive (systolic blood pressure < 90 mmHg,) blunt trauma patients has declined from 37% in 1987, to 2% in 1993. Despite a lack of data supporting efficacy in areas of severe hypotensive shock, blunt trauma, long transport times, and pelvic fractures, states continue to expend resources on the MAST suit. It is for this reason that we believe that the clinical use of the MAST suit should be based upon medical control philosophy rather than legislation.

Contact dermatitis associated with capsaicin: Hunan hand syndrome.

"Hunan hand" is a contact dermatitis resulting from the direct handling of chili peppers containing capsaicin. Capsaicin also is found in an over-the-counter topical agent for treatment of postherpetic neuralgia, diabetic neuropathy, and arthritis. We present the case of a patient with capsaicin-induced dermatitis and discuss the pathophysiology, therapy, and current uses of capsaicin.

The impact of a regionalized trauma system on trauma care in San Diego County.

A review of autopsy reports on traumatic deaths in 1986 was conducted to determine the impact on trauma mortality of the regionalized trauma system instituted in San Diego County in 1984. Determination of preventable death was made by a panel of experts and compared with an identical review of traumatic deaths in 1979, five years before the institution of regionalized trauma care. Of 211 traumatic deaths reviewed from 1986, two (1%) were classified as preventable, compared with 20 of 177 (11.4%) deaths in 1979 (P less than .001). A breakdown of trauma deaths into central nervous system and noncentral nervous system categories revealed the overall decline was in large part a consequence of the decline in non-central nervous system deaths from 16 of 83 in 1979 to one of 62 in 1986 (P less than .005). The decrease in central nervous system-related preventable deaths from four of 94 in 1979 to one of 149 in 1986 (P less than .10) was not statistically significant. Although it is likely the trauma system introduced in 1984 contributed to the decline in preventable death, it is not possible to isolate this variable from other changes that occurred during the interval between studies. A review of trauma deaths over the same time interval in a community with similar demographics but without a trauma system might help determine the relative contribution of the trauma system.