Blindness as a complication of Le Fort I osteotomy for maxillary distraction

High Le Fort I osteotomy and maxillary distraction has become an accepted method for the treatment of maxillary retrusion in children and teenagers with cleft lip and palate or craniofacial anomalies. This procedure effectively corrects the dentofacial deformity in these patients. No major surgical morbidity has been reported. During the past 4 years, 94 cleft patients with maxillary hypoplasia received Le Fort I osteotomy and distraction osteogenesis at the authors' center. Two of them developed blindness after this operation. The first case was a girl with bilateral cleft lip and palate with median facial dysplasia. She received high Le Fort I osteotomy at age 12 years 4 months to correct maxillary retrusion. Right eye swelling and ecchymosis was found after surgery. The patient complained of vision loss in that eye 2 days later. Computed tomography showed subarachnoid hemorrhage and skull base hematoma. There were no atypical fractures in the orbit, pterygoid plates, sphenoid bone, and skull base. Angiogram revealed left ophthalmic and basilar artery aneurysm. The second case was a 12-year-old boy with left cleft lip and palate. He received Le Fort I osteotomy to correct maxillary retrusion. During surgery, abnormal pupil dilatation was found after the osteotomy and down-fracture of maxilla. Emergent computed tomography found no hemorrhage or atypical fractures. Examination revealed complete left optic neuropathy and partial right abducens nerve palsy with mydriasis. Magnetic resonance imaging, magnetic resonance angiography, and repeated computed tomography revealed no sign of orbital injury, vascular problem, or abnormal fractures. The cause of blindness was unknown. In both cases, a steroid was used. Maxillary distraction was continued. Recovery of meaningful visual sense did not occur after 3 and 2 years' follow-up, respectively. A review of the literature revealed five other patients who suffered from visual loss after Le Fort I osteotomy. Inadvertent skull base fractures were identified in two cases, but a cause for the blindness was not known in the others. Induced hypotension and indirect trauma may be responsible for the optic nerve injury. In none of the cases was meaningful visual sense recovered, although high-dose steroids were given. In conclusion, a total of seven cases developed blindness after Le Fort I osteotomy. Once blindness develops, the prognosis is poor. High Le Fort I osteotomy should be performed with extreme care, and perhaps the informed consent should include visual loss as a complication of the procedure.     

Green fluorescent protein-labeled recombinant fluobody for detecting the picloram herbicide

A green fluorescent protein-labeled fluobody was designed to develop a simple immunoassay method for detecting picloram herbicide in an environmental sample. The gfp gene was successfully inserted into the pSJF2 vector harboring the picloram-specific antibody fragment to yield pSJF2GFP. Picloram spiking in an environmental river sample could be indirectly detected by observing the fluorescence intensity value of the gfp-fluobody, exhibiting specific sensitivity to free picloram with an IC50 value of 50 ppb. Using the gfp-fluobody immunoassay avoids the enzyme-substrate reaction for calorimetric detection that is required in an enzyme-linked immunosorbent assay (ELISA).     

Structural basis of tropism of Escherichia coli to the bladder during urinary tract infection

The first step in the colonization of the human urinary tract by pathogenic Escherichia coli is the mannose-sensitive binding of FimH, the adhesin present at the tip of type 1 pili, to the bladder epithelium. We elucidated crystallographically the interactions of FimH with D-mannose. The unique site binding pocket occupied by D-mannose was probed using site-directed mutagenesis. All but one of the mutants examined had greatly diminished mannose-binding activity and had also lost the ability to bind human bladder cells. The binding activity of the mono-saccharide D-mannose was delineated from this of mannotriose (Man(alpha 1-3)[Man(alpha 1-6)]Man) by generating mutants that abolished D-mannose binding but retained mannotriose binding activity. Our structure/function analysis demonstrated that the binding of the monosaccharide alpha-D-mannose is the primary bladder cell receptor for uropathogenic E. coli and that this event requires a highly conserved FimH binding pocket. The residues in the FimH mannose-binding pocket were sequenced and found to be invariant in over 200 uropathogenic strains of E. coli. Only enterohaemorrhagic E. coli (EHEC) possess a sequence variation within the mannose-binding pocket of FimH, suggesting a naturally occurring mechanism of attenuation in EHEC bacteria that would prevent them from being targeted to the urinary tract.     

5 alpha-reductase-catalyzed conversion of testosterone to dihydrotestosterone is increased in prostatic adenocarcinoma cells: suppression by 15-lipoxygenase metabolites of gamma-linolenic and eicosapentaenoic acids

Although the androgens, testosterone (T) and its highly active metabolite dihydrotestosterone (DHT) play a role in the development and progression of prostate cancer, the mechanism(s) are unclear. Furthermore, 5 alpha-reductase which catalyze the conversion of T to DHT, has been a target of manipulation in the treatment of prostatic cancer, hence synthetic 5 alpha-reductase activity inhibitors have shown therapeutic promise. To demonstrate that nutrients derived from dietary sources can exert similar therapeutic promise, this study was designed using benign hyperplastic cells (BHC) and malignant tumorigenic cells (MTC) derived from Lobund-Wistar (L-W) rat model of prostatic adenocarcinoma to test the effects of gamma-linolenic acid (GLA), eicosapentaenoic acid (EPA) and their 15-lipoxygenase metabolites on cellular 5 alpha-reductase activity. Our data revealed: (i) that incubation of MTC with [3H]-T resulted in marked conversion to [3H]-DHT when compared to similar incubation with BHC; (ii) that DHT-enhanced activity of 5 alpha-reductase was inhibited 80% by 15S-hydroxyeicosatrienoic acid, the 15-lipoxygenase metabolite of GLA, when compared to 55% by 15S-hydroxyeicosapentaenoic acid, the 15-lipoxygenase metabolite of EPA; and (iii) that their precursor fatty acids, respectively, exerted moderate inhibition. Taken together, the study underscores the biological importance of 15-lipoxygenase metabolites of polyunsaturated fatty acids (PUFAs) in androgen metabolism.     

Detection of tumor mutations in the presence of excess amounts of normal DNA

Mutations are important markers in the early detection of cancer. Clinical specimens such as bodily fluid samples often contain a small percentage of mutated cells in a large background of normal cells. Thus, assays to detect mutations leading to cancer need to be highly sensitive and specific. In addition, they should be possible to carry out in an automated and high-throughput manner to allow large-scale screening. Here we describe a screening method, termed PPEM (PNA-directed PCR, primer extension, MALDI-TOF), that addresses these needs more effectively than do existing methods. DNA samples are first amplified using peptide nucleic acid (PNA)-directed PCR clamping reactions in which mutated DNA is preferentially enriched. The PCR-amplified DNA fragments are then sequenced through primer extension to generate diagnostic products. Finally, mutations are identified using matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. This method can detect as few as 3 copies of mutant alleles in the presence of a 10,000-fold excess of normal alleles in a robust and specific manner. In addition, the method can be adapted for simultaneous detection of multiple mutations and is amenable to high-throughput automation.     

The Arabidopsis SKU5 gene encodes an extracellular glycosyl phosphatidylinositol-anchored glycoprotein involved in directional root growth

To investigate how roots respond to directional cues, we characterized a T-DNA-tagged Arabidopsis mutant named sku5 in which the roots skewed and looped away from the normal downward direction of growth on inclined agar surfaces. sku5 roots and etiolated hypocotyls were slightly shorter than normal and exhibited a counterclockwise (left-handed) axial rotation bias. The surface-dependent skewing phenotype disappeared when the roots penetrated the agar surface, but the axial rotation defect persisted, revealing that these two directional growth processes are separable. The SKU5 gene belongs to a 19-member gene family designated SKS (SKU5 Similar) that is related structurally to the multiple-copper oxidases ascorbate oxidase and laccase. However, the SKS proteins lack several of the conserved copper binding motifs characteristic of copper oxidases, and no enzymatic function could be assigned to the SKU5 protein. Analysis of plants expressing SKU5 reporter constructs and protein gel blot analysis showed that SKU5 was expressed most strongly in expanding tissues. SKU5 was glycosylated and modified by glycosyl phosphatidylinositol and localized to both the plasma membrane and the cell wall. Our observations suggest that SKU5 affects two directional growth processes, possibly by participating in cell wall expansion.     

A new family of highly variable proteins in the Chlamydophila pneumoniae genome

Chlamydiaceae are obligate intracellular bacterial pathogens characterized by a wide range of vertebrate host, tissue tropism and spectrum of diseases. To get insights into the biological mechanisms involved in these differences, we have put forward a computational and experimental procedure to identify the genome recombination hotspots, as frequent sequence variation allows rapid adaptation to environmental changes. We find a larger potential for recombination in Chlamydophila pneumoniae genomes as compared with Chlamydia trachomatis or Chlamydia muridarum. Such potential is mostly concentrated in a family of seven previously uncharacterized species-specific elements that we named ppp for C.pneumoniae polymorphic protein genes, which have the potential to vary by homologous recombination and slipped-mispair. Experimentally, we show that these sequences are indeed highly polymorphic among a collection of nine C.pneumoniae strains of very diverse geographical and pathological origins, mainly by slippage of a poly(C) tract. We also show that most elements are transcribed during infection. In silico analyses suggest that Ppps correspond to outer membrane proteins. Given their species specificity, their putative location in the outer membrane and their extreme polymorphism, Ppps are most likely to be important in the pathogenesis of C.pneumoniae and could represent targets for future vaccine development.