Effect of Transverse Versus Longitudinal Incisions on Anterior Knee Pain After Tibial Nailing (TRAVEL): A Multicenter Randomized Trial with 1-Year Follow-up.

BACKGROUND: Anterior knee pain is common after tibial nailing, and its origin is poorly understood. Literature suggests it may be related to infrapatellar nerve injury. The aim of this study was to compare the effect of a transverse (potentially infrapatellar nerve-sparing) incision versus a longitudinal incision for the insertion of a tibial nail with respect to anterior knee pain. METHODS: Patients with a tibial shaft fracture undergoing treatment with use of an intramedullary nail were randomized to a transverse incision (n = 68) or longitudinal incision (n = 68) in multiple centers. The primary outcome measure was kneeling pain based on a numeric rating scale (NRS). Secondary outcome measures included knee pain during daily activities, functional outcome (Short Musculoskeletal Function Assessment and Lower Extremity Functional Scale), quality of life (EuroQol-5 Dimensions), activity resumption, complications, reoperations, and costs within 1 year after trauma. RESULTS: At 12 months, the estimated marginal mean for kneeling pain was 2.4 (95% confidence interval [CI], 1.6 to 3.2) in the transverse incision group and 3.7 (95% CI, 3.0 to 4.5) in the longitudinal incision group. Regression analysis showed no significant difference between the groups over time. Knee pain scores for daily activities, functional outcome scores, and quality of life were also comparable between the groups. Signs of infrapatellar nerve injury were found less often after a transverse incision (18% versus 54%; p < 0.001). The median total (direct and indirect) costs per patient were €10,468 in the transverse incision group and €11,066 in the longitudinal incision group. Loss of productivity accounted for 67% and 52% of the total costs in the 2 groups, respectively. CONCLUSIONS: A transverse incision reduces injury to the infrapatellar nerve, but anterior knee pain scores and function are comparable after use of a transverse or longitudinal incision for tibial nail insertion. LEVEL OF EVIDENCE: Therapeutic Level I . See Instructions for Authors for a complete description of levels of evidence.

Measurement properties of Patient-Reported Outcome Measures in patients with a tibial shaft fracture; validation study alongside the multicenter TRAVEL study.

The aim of this study was to evaluate the measurement properties of the Short Musculoskeletal Function Assessment (SMFA) and Lower Extremity Functional Scale (LEFS) in patients who sustained a tibial shaft fracture, by comparing them with the scores of a general health-related quality of life instrument scale (i.e., EuroQoL-5D). Data of 136 patients participating in a multicenter randomized controlled trial comparing incisions for intramedullary nail entry in adults with a tibial shaft fracture were used. Patients completed the SMFA, LEFS, EQ-5D and an anchor question at 2 and 6 weeks, and at 3, 6 and 12 months. Reliability (internal consistency), construct validity, responsiveness (longitudinal validity), floor and ceiling effects, minimal important change (MIC), and smallest detectable change (SDC) were determined. The SMFA and LEFS (sub)scales showed adequate internal consistency (0.84<α<0.94). Construct and longitudinal validity were also adequate (correctly predicted hypotheses between 83%-100%). Floor effects were not present. Ceiling effects were present at 12 months for the SMFA lower extremity dysfunction and bother subscales (22% and 19%, respectively) and the LEFS (19%). MICs could not be determined with the available data. The SDC was 13.84 points for the SMFA and 38.74 points for the LEFS. This study confirms that the SMFA and LEFS are reliable, valid, and responsive instruments for monitoring functional limitation in patients after sustaining a tibia shaft fracture during at least the first six months post-injury. An anchor-based MIC for the SMFA remains to be determined.

Small bowel gastrointestinal stromal tumor presenting with gastrointestinal bleeding in patient with type 1 Neurofibromatosis: Management and laparoscopic treatment. Case report and review of the literature.

INTRODUCTION AND IMPORTANCE: Gastrointestinal stromal tumor (GIST) is the most common mesenchymal neoplasm of the gastrointestinal tract. It may be asymptomatic; nevertheless, gastrointestinal bleeding is the most frequent symptom, due to mucosal erosion. Its poor lymph node metastatic spread makes GIST often suitable of minimally invasive surgical approach. The importance of this study is to increase the awareness among physicians about this condition in particular scenarios as in our case and to stress the role of laparoscopic surgery. CASE PRESENTATION: A 74-year-old female patient presented to the emergency department with hematemesis, followed by haematochezia and melena. The patient had a medical history of type 1 Neurofibromatosis (NF1). She underwent, after CT scan, esophagogastroduodenoscopy, and endoscopic haemostasis. Finally, we performed a laparoscopic resection of a mass of the first jejunal loop. The postoperative period was predominantly uneventful. Pathological examination confirmed a low-risk GIST. CLINICAL DISCUSSION: Proximal jejunal GIST may cause an upper and lower gastrointestinal bleeding. A multidisciplinary team approach is mandatory for the correct management of this disease and its complications (bleeding). GISTs are indicated as the most commonly gastrointestinal NF1 associated tumours. In case of localised and resectable GIST surgical treatment is the mainstay and laparoscopic surgery is a valid alternative. CONCLUSION: In case of abdominal bleeding mass in a NF1 patient, it is important to keep in mind the well-known association between NF1 and GIST to facilitate the diagnosis and to quickly perform the appropriate treatment. Laparoscopic approach is safe and effective if the oncological radicality is respected.

Anterior knee pain and functional outcome following different surgical techniques for tibial nailing: a systematic review.

PURPOSE: The aim of this systematic review was to compare knee pain and function after tibial nail insertion through an infrapatellar, semi-extended and suprapatellar technique. METHODS: A search was carried out to identify articles with an exact description of the method used for insertion of the tibial nail and description of the outcome parameters (knee pain or function). Data on study design, population, rate and severity of anterior knee pain and function scores were extracted. Pooled rates and scores were calculated. RESULTS: 67 studies with 3,499 patients were included. The pooled rate of patients with anterior knee pain was 38% (95% CI 32-44) after nail insertion through an infrapatellar approach and 10% (95% CI 1-26) after insertion through a suprapatellar approach. Pooled analysis was not possible for the semi-extended technique. Knee pain scores as measured by visual analogue score (0-10) ranged from 0.2 (95% CI - 0.1-0.5) for general knee pain to 3.7 (95% CI 1.3-6.1) for pain during kneeling. Pooled estimates for the Lysholm score were 87 points (range 77-97) for the infrapatellar technique and 85 points (range 82-85) for the suprapatellar technique. Iowa Knee scores were 94 (range 86-96) and Anterior Knee Pain Scale scores were 76 (range 75-80) after infrapatellar nail insertion. DISCUSSION: Depending on the technique used, the proportion of patients with knee pain after tibial nailing varied between 10 and 38%. The actual measured knee pain scores were, however, surprisingly low. Knee function was good for both the infra- and suprapatellar technique.

Health care and productivity costs for isolated tibia shaft fracture admissions in The Netherlands.

The aim of this study was to provide a detailed overview of age and gender specific health care costs and costs due to lost productivity for hospital admitted patients with an isolated tibia shaft fracture in The Netherlands between 2008 and 2012. Injury cases and length of hospital stay were extracted from the National Medical Registration. Information on extramural health care and work absence were retrieved from a patient follow-up survey on health care use. Medical costs included ambulance care, in- hospital care, general practitioner care, home care, physical therapy, and rehabilitation/nursing care. An incidence-based cost model was applied to calculate direct health care costs and lost productivity in 2012. Total direct health care costs for all patients admitted with a tibia shaft fracture (n = 1,635) were €13.6 million. Costs for productivity loss were € 23.0 million. Total costs (direct health care and lost productivity) per patient were highest for men aged 40-49 years mainly due to lost productivity, and for women aged > 80 years, due to high direct medical costs.

Epidemiologic trends for isolated tibia shaft fracture admissions in The Netherlands between 1991 and 2012.

INTRODUCTION: Population-based knowledge on the occurrence of specific injuries is essential for the allocation of health care services, optimization of preventive measures, and research purposes. Therefore, the aim of this study was to examine long-term nation-based trends in the incidence rate, trauma mechanism, hospital length of stay (HLOS), treatment, and outcome of hospital-admitted patients with an isolated tibia shaft fracture between 1991 and 2012 in The Netherlands. METHODS: All hospital-admitted patients in The Netherlands between 1991 and 2012 with an isolated tibia shaft fracture were included. Age and gender-standardized incidence rates were calculated for each year. Data were extracted from the National Medical Registration. RESULTS: The incidence rate for men decreased to 13.8/100,000 person years (py). For women the incidence rate remained stable with 7.2/100,000 py. Incidence showed a peak for adolescent men (15-19 years), and increased in both genders from 65 years onwards. Since 1993 the mean HLOS for isolated tibia fractures reduced from 10.8 to 5.4 days. Mean HLOS increased with age. Mean years lived with disability (YLD) was 4.5 years, declined linearly with age, and showed no gender effect. CONCLUSIONS: In 22 years, the incidence rate of hospital admitted patients with an isolated tibia shaft fracture in The Netherlands dropped with 12%, which was mainly attributable to a 15% decline among men. Incidence rate, trauma mechanism, and HLOS were age and gender related. HLOS also reduced over time. Operation rate and YLD were only age related.

An infrapatellar nerve block reduces knee pain in patients with chronic anterior knee pain after tibial nailing: a randomized, placebo-controlled trial in 34 patients.

Background and purpose - Anterior knee pain is common after tibial nailing. Its origin is poorly understood. Injury of the infrapatellar nerve is a possible cause. In this randomized controlled trial we compared changes in knee pain after an infrapatellar nerve block with lidocaine or placebo in patients with persistent knee pain after tibial nailing. Patients and methods - Patients with chronic knee pain after tibial nailing were randomized to an infrapatellar nerve block with 5 ml 2% lidocaine or placebo (sodium chloride 0.9%), after which they performed 8 daily activities. Before and after these activities, pain was recorded using a numeric rating scale (NRS; 0-10). Primary endpoint was the change in pain during kneeling after the infrapatellar nerve block. Secondary outcomes were changes in pain after the nerve block during the other activities. Results - 34 patients (age 18-62 years) were equally randomized. A significant reduction of the NRS for kneeling pain with an infrapatellar nerve block with lidocaine was found compared with placebo (-4.5 [range -10 to -1] versus -1 [-9 to 2]; p = 0.002). There were no differences between the treatments for the NRS values for pain during other activities. Interpretation - Compared with placebo, an infrapatellar nerve block with lidocaine was more effective in reducing pain during kneeling in patients with chronic knee pain after tibial nailing. Our findings support the contention that kneeling pain after tibial nailing is a peripheral nerve-related problem.

Laparoscopic treatment of incisional and primary ventral hernia in morbidly obese patients with a BMI over 35.

INTRODUCTION: Incisional and ventral hernias are common surgical indications. Their management is associated with significant complications and recurrences in open surgery (15-25%). Since laparoscopy has become a standard in bariatric surgery, there has been a natural trend to treat obese patients with parietal wall defects laparoscopically. The aim of our study was to evaluate the feasibility and the results of the laparoscopic management of parietal wall defects in patients with a BMI >35. MATERIALS AND METHODS: A series of 79 patients were included. Data were acquired prospectively and analyzed retrospectively. The surgical procedure was standardized: 3 ports, mesh type (Parietex™ Composite mesh, Covidien, France), fixation with non-absorbable transfascial sutures, and tackers. Complications were evaluated. RESULTS: Out of 79 patients (29 men, 50 women), 43 had umbilical and 36 had ventral hernias. Mean age was 52.4 years, and mean BMI was 40.83 kg/m(2). Mean postoperative hospital stay was 2 days. Postoperative pain evaluated by visual analog scale was 2.86. No intraoperative complications or deaths occurred. Seven postoperative complications occurred (8.86%): two parietal wall hematomas treated by radiological embolization, two significant cases of postoperative pain, one postoperative obstruction, one spontaneously resolved respiratory failure, and one early (day 1) parietal wall defect with immediate reoperation. Postoperative seroma rate was 26.58% (21 patients, all of whom were treated conservatively). Postoperative follow-up was 18.10 months (1-84 months), and recurrence rate was 3.8% (3 patients). DISCUSSION: This study confirms the feasibility and safety of the laparoscopic approach for ventral hernias in morbidly obese patients. Recurrence rates (3.8%) appeared lower than the ones observed in the literature (15-25%). Postoperative hemorrhage and port-site hernia are specific complications of this approach. Postoperative hospital stay is low (2 days) as compared to open surgery. Laparoscopic management of parietal wall defects should be considered a standard option in morbidly obese patients.

[A woman with a chronic wound on the clavicle]. / Een vrouw met een chronische wond op de clavicula.

A 49-year-old woman, who was treated several times with radiotherapy for breast cancer, developed a chronic wound on her right clavicle.

Immune cell regulation and cardiovascular effects of sphingosine 1-phosphate receptor agonists in rodents are mediated via distinct receptor subtypes.

Sphingosine 1-phosphate (S1P) is a bioactive lysolipid with pleiotropic functions mediated through a family of G protein-coupled receptors, S1P(1,2,3,4,5). Physiological effects of S1P receptor agonists include regulation of cardiovascular function and immunosuppression via redistribution of lymphocytes from blood to secondary lymphoid organs. The phosphorylated metabolite of the immunosuppressant agent FTY720 (2-amino-2-(2-[4-octylphenyl]ethyl)-1,3-propanediol) and other phosphonate analogs with differential receptor selectivity were investigated. No significant species differences in compound potency or rank order of activity on receptors cloned from human, murine, and rat sources were observed. All synthetic analogs were high-affinity agonists on S1P(1), with IC(50) values for ligand binding between 0.3 and 14 nM. The correlation between S1P(1) receptor activation and the ED(50) for lymphocyte reduction was highly significant (p < 0.001) and lower for the other receptors. In contrast to S1P(1)-mediated effects on lymphocyte recirculation, three lines of evidence link S1P(3) receptor activity with acute toxicity and cardiovascular regulation: compound potency on S1P(3) correlated with toxicity and bradycardia; the shift in potency of phosphorylated-FTY720 for inducing lymphopenia versus bradycardia and hypertension was consistent with affinity for S1P(1) relative to S1P(3); and toxicity, bradycardia, and hypertension were absent in S1P(3)(-/-) mice. Blood pressure effects of agonists in anesthetized rats were complex, whereas hypertension was the predominant effect in conscious rats and mice. Immunolocalization of S1P(3) in rodent heart revealed abundant expression on myocytes and perivascular smooth muscle cells consistent with regulation of bradycardia and hypertension, whereas S1P(1) expression was restricted to the vascular endothelium.

Sphingosine-1-phosphate phosphatases.

Sphingosine-1-phosphate is a potent proliferative, survival, and morphogenetic factor, acting as an extracellular ligand for the EDG family of G-protein-coupled receptors and possibly intracellularly through as yet, unidentified targets. It is produced within most, if not all cells by phosphorylation of sphingosine, and is an abundant serum lipid that is released from activated platelets. Sphingosine and sphingosine-1-phosphate are in dynamic equilibrium with each other due to the activities of sphingosine kinase and sphingosine-1-phosphate phosphatase (SPPase). Several SPPase genes have now been cloned, first from yeast and more recently from mammalian cells. By sequence homology, these enzymes can be classified as a subset of membrane bound, Type 2 lipid phosphohydrolases that contain conserved residues within three domains predicted to be at the active site of the enzyme. Outside of the consensus motif, there is very little homology between SPPases and the other type 2 lipid phosphohydrolases in the LPP/PAP family. Type 2 phosphatase activity is Mg+-independent and insensitive to N-ethylmaleimide, and substrate specificity is broad for LPP enzymes, whereas SPPases are highly selective for sphingolipid substrates. SPPase activity in yeast and mammalian cells regulates intracellular sphingosine-1-phosphate levels, and also alters the levels of sphingosine and ceramide, two other signaling molecules that often oppose the actions of sphingosine-1-phosphate. Thus, loss of SPPase in yeast results in high sphingosine-1-phosphate levels and cells are more resistant to stress, and in mammalian cells, overexpression of SPPase elevates ceramide levels and provokes apoptosis.

The effect of the erg26-1 mutation on the regulation of lipid metabolism in Saccharomyces cerevisiae.

A temperature-sensitive Saccharomyces cerevisiae mutant harboring a lesion in the ERG26 gene has been isolated. ERG26 encodes 4alpha-carboxysterol-C3 dehydrogenase, one of three enzymatic activities required for the conversion of 4,4-dimethylzymosterol to zymosterol. Gas chromatography/mass spectrometry analyses of sterols in this mutant, designated erg26-1, revealed the aberrant accumulation of a 4-methyl-4-carboxy zymosterol intermediate, as well as a novel 4-carboxysterol. Neutral lipid radiolabeling studies showed that erg26-1 cells also harbored defects in the rate of biosynthesis and steady-state levels of mono-, di-, and triglycerides. Phospholipid radiolabeling studies showed defects in the rate of biosynthesis of both phosphatidic acid and phosphatidylinositol. Biochemical studies revealed that microsomes isolated from erg26-1 cells contained greatly reduced 4alpha-carboxysterol-C3 dehydrogenase activity when compared with microsomes from wild type cells. Previous studies have shown that loss of function mutations in either of the fatty acid elongase genes SUR4/ELO3 or FEN1/GNS1/ELO2 can "bypass" the essentiality of certain ERG genes (Ladeveze, V., Marcireau, C., Delourme, D., and Karst, F. (1993) Lipids 28, 907-912; Silve, S., Leplatois, P., Josse, A., Dupuy, P. H., Lanau, C., Kaghad, M., Dhers, C., Picard, C., Rahier, A., Taton, M., Le Fur, G., Caput, D., Ferrara, P., and Loison, G. (1996) Mol. Cell. Biol. 16, 2719-2727). Studies presented here have shown that this sphingolipid-dependent "bypass" mechanism did not suppress the essential requirement for zymosterol biosynthesis. However, studies aimed at understanding the underlying physiology behind the temperature-sensitive growth defect of erg26-1 cells showed that the addition of several antifungal compounds to the growth media of erg26-1 cells could suppress the temperature-sensitive growth defect. Fluorescence microscopic analysis showed that GFP-Erg26p and GFP-Erg27p fusion proteins were localized to the endoplasmic reticulum. Two-hybrid analysis indicated that Erg25p, Erg26p, and Erg27p, which are required for the biosynthesis of zymosterol, form a complex within the cell.

Molecular cloning and characterization of a lipid phosphohydrolase that degrades sphingosine-1- phosphate and induces cell death.

Sphingosine and sphingosine-1-phosphate (SPP) are interconvertible sphingolipid metabolites with opposing effects on cell growth and apoptosis. Based on sequence homology with LBP1, a lipid phosphohydrolase that regulates the levels of phosphorylated sphingoid bases in yeast, we report here the cloning, identification, and characterization of a mammalian SPP phosphatase (mSPP1). This hydrophobic enzyme, which contains the type 2 lipid phosphohydrolase conserved sequence motif, shows substrate specificity for SPP. Partially purified Myc-tagged mSPP1 was also highly active at dephosphorylating SPP. When expressed in yeast, mSPP1 can partially substitute for the function of LBP1. Membrane fractions from human embryonic kidney HEK293 cells transfected with mSPP1 markedly degraded SPP but not lysophosphatidic acid, phosphatidic acid, or ceramide-1-phosphate. Enforced expression of mSPP1 in NIH 3T3 fibroblasts not only decreased SPP and enhanced ceramide levels, it also markedly diminished survival and induced the characteristic traits of apoptosis. Collectively, our results suggest that SPP phosphohydrolase may regulate the dynamic balance between sphingolipid metabolite levels in mammalian cells and consequently influence cell fate.

Characterization of a novel sphingosine 1-phosphate receptor, Edg-8.

Three G protein-coupled receptors (Edg-1, Edg-3, and Edg-5) for the lysolipid phosphoric acid mediator sphingosine 1-phosphate have been described by molecular cloning. Using a similar sequence that we found in the expressed sequence tag data base, we cloned and characterized of a fourth, high affinity, rat brain sphingosine 1-phosphate receptor, Edg-8. When HEK293T cells were co-transfected with Edg-8 and G protein DNAs, prepared membranes showed sphingosine 1- phosphate-dependent increases in [(35)S]guanosine 5'-(3-O-thio)triphosphate binding with an EC(50) of 90 nm. In a rat hepatoma Rh7777 cell line that exhibits modest endogenous responses to sphingosine 1-phosphate, this lipid mediator inhibited forskolin-driven rises in cAMP by greater than 90% when the cells were transfected with Edg-8 DNA (IC(50) 0.7 nm). This response is blocked fully by prior treatment of cultures with pertussis toxin, thus implicating signaling through G(i/o)alpha proteins. Furthermore, Xenopus oocytes exhibit a calcium response to sphingosine 1-phosphate after injection of Edg-8 mRNA, but only when oocytes are co-injected with chimeric G(q/i)alpha protein mRNA. Membranes from HEK293T and Rh7777 cell cultures expressing Edg-8 exhibited high affinity (K(D) = 2 nm) binding for radiolabeled sphingosine 1-phosphate. Rat Edg-8 RNA is expressed in spleen and throughout adult rat brain where in situ hybridization revealed it to be associated with white matter. Together our data demonstrate that Edg-8 is a high affinity sphingosine 1-phosphate receptor that couples to G(i/o)alpha proteins and is expressed predominantly by oligodendrocytes and/or fibrous astrocytes in the rat brain.

Inhibition of fungal sphingolipid biosynthesis by rustmicin, galbonolide B and their new 21-hydroxy analogs.

The mode of action of the known antifungal macrolides rustmicin (1) and galbonolide B (2) has been determined to be the inhibition of sphingolipid biosynthesis. A large scale fermentation and isolation process was developed for production of large quantities of rustmicin. New 21-hydroxy derivatives of both compounds were isolated from pilot scale fermentations and were also produced by biotransformation of rustmicin and galbonolide B.

Rustmicin, a potent antifungal agent, inhibits sphingolipid synthesis at inositol phosphoceramide synthase.

Rustmicin is a 14-membered macrolide previously identified as an inhibitor of plant pathogenic fungi by a mechanism that was not defined. We discovered that rustmicin inhibits inositol phosphoceramide synthase, resulting in the accumulation of ceramide and the loss of all of the complex sphingolipids. Rustmicin has potent fungicidal activity against clinically important human pathogens that is correlated with its sphingolipid inhibition. It is especially potent against Cryptococcus neoformans, where it inhibits growth and sphingolipid synthesis at concentrations <1 ng/ml and inhibits the enzyme with an IC50 of 70 pM. This inhibition of the membrane-bound enzyme is reversible; moreover, rustmicin is nearly equipotent against the solubilized enzyme. Rustmicin was efficacious in a mouse model for cryptococcosis, but it was less active than predicted from its in vitro potency against this pathogen. Stability and drug efflux were identified as two factors limiting rustmicin's activity. In the presence of serum, rustmicin rapidly epimerizes at the C-2 position and is converted to a gamma-lactone, a product that is devoid of activity. Rustmicin was also found to be a remarkably good substrate for the Saccharomyces cerevisiae multidrug efflux pump encoded by PDR5.

Sphingoid base 1-phosphate phosphatase: a key regulator of sphingolipid metabolism and stress response.

The sphingolipid metabolites ceramide and sphingosine-1-phosphate are second messengers with opposing roles in mammalian cell growth arrest and survival; their relative cellular level has been proposed to be a rheostat that determines the fate of cells. This report demonstrates that this rheostat is an evolutionarily conserved stress-regulatory mechanism that influences growth and survival of yeast. Although the role of sphingosine-1-phosphate in yeast was not previously examined, accumulation of ceramide has been shown to induce G1 arrest and cell death. We now have identified a gene in Saccharomyces cerevisiae, LBP1, that regulates the levels of phosphorylated sphingoid bases and ceramide. LBP1 was cloned from a yeast mutant that accumulated phosphorylated long-chain sphingoid bases and diverted sphingoid base intermediates from sphingolipid pathways to glycerophospholipid biosynthesis. LBP1 and its homolog, LBP2, encode very hydrophobic proteins that contain a novel-conserved sequence motif for lipid phosphatases, and both have long-chain sphingoid base phosphate phosphatase activity. In vitro characterization of Lbp1p shows that this phosphatase is Mg2+-independent with high specificity for phosphorylated long-chain bases, phytosphingosine and sphingosine. The deletion of LBP1 results in the accumulation of phosphorylated long-chain sphingoid bases and reduced ceramide levels. Moreover, deletion of LBP1 and LBP2 results in dramatically enhanced survival upon severe heat shock. Thus, these phosphatases play a previously unappreciated role in regulating ceramide and phosphorylated sphingoid base levels in yeast, and they modulate stress responses through sphingolipid metabolites in a manner that is reminiscent of their effects on mammalian cells.

ELO2 and ELO3, homologues of the Saccharomyces cerevisiae ELO1 gene, function in fatty acid elongation and are required for sphingolipid formation.

ELO2 and ELO3 were identified from the Saccharomyces cerevisiae genome data base as homologues of ELO1, a gene involved in the elongation of the fatty acid 14:0 to 16:0. Mutations in these genes have previously been shown to produce pleiotropic effects involving a number of membrane functions. The simultaneous disruption of ELO2 and ELO3 has also been shown to produce synthetic lethality, indicating that they have related and/or overlapping functions. Gas chromatography and gas chromatography/mass spectroscopy analyses reveal that null mutations of ELO2 and ELO3 produce defects in the formation of very long chain fatty acids. Analysis of the null mutants indicates that these genes encode components of the membrane-bound fatty acid elongation systems that produce the 26-carbon very long chain fatty acids that are precursors for ceramide and sphingolipids. Elo2p appears to be involved in the elongation of fatty acids up to 24 carbons. It appears to have the highest affinity for substrates with chain lengths less than 22 carbons. Elo3p apparently has a broader substrate specificity and is essential for the conversion of 24-carbon acids to 26-carbon species. Disruption of either gene reduces cellular sphingolipid levels and results in the accumulation of the long chain base, phytosphingosine. Null mutations in ELO3 result in accumulation of labeled precursors into inositol phosphoceramide, with little labeling in the more complex mannosylated sphingolipids, whereas disruption of ELO2 results in reduced levels of all sphingolipids.