Survival after massive hydroxychloroquine overdose.

Hydroxychloroquine overdose is infrequently reported and the majority of recommendations come from the greater experience with chloroquine poisoning. We report two cases of massive hydroxychloroquine poisoning (20 g in each case), both of which received advanced cardiac life support and a treatment regimen consisting of sodium bicarbonate, adrenaline and potassium. Both these patients survived beyond their initial rapid deterioration and cardiovascular collapse to be discharged from hospital without sequelae. These patients had the highest reported non-lethal serum concentrations (13.8 and 26.0 mg/l). They both demonstrated rapid recovery from a pre-arrest condition, following aggressive correction of electrolyte and pH disturbance and rapid distribution of the drug to peripheral tissues.

Cloning of matrix Gla protein in a marine cartilaginous fish, Prionace glauca: preferential protein accumulation in skeletal and vascular systems.

Matrix Gla protein (MGP) belongs to the family of vitamin K dependent, Gla containing proteins and, in mammals, birds and Xenopus, its mRNA has been previously detected in bone, cartilage and soft tissue extracts, while the accumulation of the protein was found mainly in calcified tissues. More recently, the MGP gene expression was also studied in marine teleost fish where it was found to be associated with chondrocytes, smooth muscle and endothelial cells. To date no information is available on the sites of MGP expression or accumulation in cartilaginous fishes that diverged from osteichthyans, a group that includes mammals, over 400 million years ago. The main objectives of this work were to study the sites of MGP gene expression and protein accumulation by means of in situ hybridization and immunohistochemistry. MGP mRNA and protein were localized as expected not only in cartilage from branchial arches and vertebra but also in the endothelia of the vascular system as well as in the tubular renal endothelium. The accumulation of MGP in non mineralized soft tissues was unexpected and suggests differences in localization or regulation of this protein in shark soft tissues compared to tetrapods and teleosts. Our results also corroborate the hypothesis that in Prionace glauca, as previously shown in mammals, the MGP protein probably also acts as a calcification inhibitor, protecting soft tissues from abnormal and ectopic calcification.

Osteocalcin and matrix GLA protein in developing teleost teeth: identification of sites of mRNA and protein accumulation at single cell resolution.

In this study, the tissue distribution and accumulation of osteocalcin or bone Gla protein (BGP) and matrix Gla protein (MGP) were determined during tooth development in a teleost fish, Argyrosomus regius. In this species, the presence of BGP and MGP mRNA in teeth was revealed by in situ hybridization. mRNA for BGP was detected in the odontoblasts as well as in its cytoplasmic processes emerging through dentinal tubules, while mRNA for MGP was expressed in the enamel portion within the apical portion of the elongated cell bodies of enameloblasts, adjacent to the root of the teeth as well as in cells within the pulpal space. Immunolocalization of BGP and MGP demonstrated that these proteins accumulate mainly in the mineralized dentin or in enameloblastic processes, confirming in situ hybridization results. In this study, we examined for the first time the localization of both BGP and MGP gene expression and protein accumulation within the different regions of the vertebrate tooth. We clearly demonstrated that although the overall pattern of BGP and MGP gene expression and protein accumulation in A. regius teeth was in general agreement to what is known for other vertebrates such as rats or rodents, our study provided novel information and highlighted some species-differences between fish and higher vertebrates.

Supplemental jet ventilation in a case of ARDS complicated by bronchopleural fistulae.

OBJECTIVE: To present a case of unusual ventilatory strategy in a 17 year old girl with the acute respiratory distress syndrome (ARDS) complicated by bilateral bronchopleural fistulae. METHODS: The patient was ventilated with a combination of conventional pressure control ventilation (PCV) and high frequency jet ventilation (HFJV) for 133 and 110 days, respectively. RESULTS: Despite prolonged hypoxia, extensive barotrauma and nosocomial infections, she survived without significant impairment of respiratory function. Two years later she was healthy and independent with only mildly reduced respiratory reserve. CONCLUSIONS: The combination of PCV and HFJV was beneficial in this case of ARDS complicated by bronchopleural fistulae. The case also highlights the utility of HFJV in the desperately hypoxic patient with extensive airway disruption.

Characterization of osteocalcin (BGP) and matrix Gla protein (MGP) fish specific antibodies: validation for immunodetection studies in lower vertebrates.

In fish species the basic mechanisms of bone development and bone remodeling are not fully understood. The classification of bone tissue in teleosts as cellular or acellular and the presence of transitional states between bone and cartilage and the finding of different types of cartilage in teleosts not previously recognized in higher vertebrates emphasizes the need for a study on the accumulation of the Gla-containing proteins MGP and BGP at the cellular level. In the present study, polyclonal antibodies developed against BGP and MGP from A. regius (a local marine teleost fish) and against MGP from G. galeus (a Pacific Ocean shark), were tested by Western blot for their specificity against BGP and MGP from several other species of teleost fish and shark. For this purpose we extracted and purified both proteins from various marine and freshwater teleosts, identified them by N-terminal amino acid sequence analysis and confirmed the presence of gamma-carboxylation in the proteins with the use of a stain specific for Gla residues. Each antibody recognized either BGP or MGP with no cross-reaction between proteins detected. All purified fish BGPs and MGPs tested were shown to be specifically recognized, thus validating the use of these antibodies for further studies.

Purification of matrix Gla protein from a marine teleost fish, Argyrosomus regius: calcified cartilage and not bone as the primary site of MGP accumulation in fish.

Matrix Gla protein (MGP) belongs to the family of vitamin K-dependent, Gla-containing proteins, and in mammals, birds, and Xenopus, its mRNA was previously detected in extracts of bone, cartilage, and soft tissues (mainly heart and kidney), whereas the protein was found to accumulate mainly in bone. However, at that time, it was not evaluated if this accumulation originated from protein synthesized in cartilage or in bone cells because both coexist in skeletal structures of higher vertebrates and Xenopus. Later reports showed that MGP also accumulated in costal calcified cartilage as well as at sites of heart valves and arterial calcification. Interestingly, MGP was also found to accumulate in vertebra of shark, a cartilaginous fish. However, to date, no information is available on sites of MGP expression or accumulation in teleost fishes, the ancestors of terrestrial vertebrates, who have in their skeleton mineralized structures with both bone and calcified cartilage. To analyze MGP structure and function in bony fish, MGP was acid-extracted from the mineralized matrix of either bone tissue (vertebra) or calcified cartilage (branchial arches) from the bony fish, Argyrosomus regius, separated from the mineral phase by dialysis, and purified by Sephacryl S-100 chromatography. No MGP was recovered from bone tissue, whereas a protein peak corresponding to the MGP position in this type of gel filtration was obtained from an extract of branchial arches, rich in calcified cartilage. MGP was identified by N-terminal amino acid sequence analysis, and the resulting protein sequence was used to design specific oligonucleotides suitable to amplify the corresponding DNA by a mixture of reverse transcription-polymerase chain reaction (RT-PCR) and 5'rapid amplification of cDNA (RACE)-PCR. In parallel, ArBGP (bone Gla protein, osteocalcin) was also identified in the same fish, and its complementary DNA cloned by an identical procedure. Tissue distribution/accumulation was analyzed by Northern blot, in situ hybridization, and immunohistochemistry. In mineralized tissues, the MGP gene was predominantly expressed in cartilage from branchial arches, with no expression detected in the different types of bone analyzed, whereas BGP mRNA was located in bone tissue as expected. Accordingly, the MGP protein was found to accumulate, by immunohistochemical analysis, mainly in the extracellular matrix of calcified cartilage. In soft tissues, MGP mRNA was mainly expressed in heart but in situ hybridization, indicated that cells expressing the MGP gene were located in the bulbus arteriosus and aortic wall, rich in smooth muscle and endothelial cells, whereas no expression was detected in the striated muscle myocardial fibers of the ventricle. These results show that in marine teleost fish, as in mammals, the MGP gene is expressed in cartilage, heart, and kidney tissues, but in contrast with results obtained in Xenopus and higher vertebrates, the protein does not accumulate in vertebra of non-osteocytic teleost fish, but only in calcified cartilage. In addition, our results also indicate that the presence of MGP mRNA in heart tissue is due, at least in fish, to the expression of the MGP gene in only two specific cell types, smooth muscle and endothelial cells, whereas no expression was found in the striated muscle fibers of the ventricle. In light of these results and recent information on expression of MGP gene in these same cell types in mammalian aorta, it is likely that the levels of MGP mRNA previously detected in Xenopus, birds, and mammalian heart tissue may be restricted to regions rich in smooth muscle and endothelial cells. Our results also emphasize the need to re-evaluate which cell types are involved in MGP gene expression in other soft tissues and bring further evidence that fish are a valuable model system to study MGP gene expression and regulation.

Cloning and characterization of the cDNA and gene encoding Xenopus laevis osteocalcin.

A full length Xenopus laevis osteocalcin (bone Gla protein, BGP) has been cloned by a combination of reverse transcription and amplification by the polymerase chain reaction, sequenced, and found to encode a polypeptide with 101 amino acid residues, including a 52-residue prepro-region and a 49-residue mature protein. The N-terminal region of the mature Xenopus BGP (xBGP), as deduced from the cDNA, is in full agreement with the sequence of the BGP previously purified from Xenopus long bones. This cDNA was used to clone the xBGP gene and its promoter region. The xBGP gene spans 3727 bp from the site of transcription initiation corresponding to the 5'end of the cDNA to the site of insertion of the poly-A(+) tail, and it contains four exons. This structure is similar to the one obtained for both fish and mammalian BGP genes and indicates that the molecular organization of this gene has been conserved throughout vertebrate evolution. Also similar to other known vertebrate systems, xBGP gene expression is restricted to bone, with no signal for xBGP messenger RNA (mRNA) detected in all other tissues analyzed. The availability of the xBGP promoter will permit to analyze its regulation in a widely used non-mammalian model system for vertebrate development, taking advantage of the availability of sequences for various Xenopus steroid hormone receptors and transcription factors known to affect BGP expression in the mammalian system.

Early and late removal of the pressure bandage in brown snake envenomation: a report of two cases.

Two cases of brown snake envenomation are presented where the duration of bandage application in one patient was prolonged compared with the other patient and was associated with a reduction in the total amount of antivenom required. One patient had the bandage removed 2 hours and twenty minutes after application and required 25 units of brown snake antivenom to neutralise the defibrination coagulopathy and manage an upper gastrointestinal haemorrhage. This patient also sustained an urticarial reaction during administration of the final 5 vials of antivenom. The other patient had the bandage released more than 22 hours after its application and only required a total of 6 units of brown snake antivenom to neutralise the defibrination coagulopathy. In the latter case, there was no reaction to any of the vials of antivenom These cases suggest that bandage release could be delayed well beyond the usual recommended time to effect a reduction in peak and cumulative venom levels and antivenom requirements.

Delayed bilateral internal carotid artery thrombosis following accidental strangulation.

A 24-yr-old male presented after a fishing accident in which he was pulled underwater by a rope attached to a crayfish pot. He was winched out of the water with the rope still around his neck, sustaining serious neck injuries that ultimately led to his death. After initial resuscitation, he remained fully conscious for approximately 8 h, after which there was a rapid and sudden deterioration in his level of consciousness. The presentation, investigation, management and subsequent postmortem findings are presented and discussed.