Improved Constraints on Primordial Gravitational Waves using Planck, WMAP, and BICEP/Keck Observations through the 2018 Observing Season.

We present results from an analysis of all data taken by the BICEP2, Keck Array, and BICEP3 CMB polarization experiments up to and including the 2018 observing season. We add additional Keck Array observations at 220 GHz and BICEP3 observations at 95 GHz to the previous 95/150/220 GHz dataset. The Q/U maps now reach depths of 2.8, 2.8, and 8.8 µK_{CMB} arcmin at 95, 150, and 220 GHz, respectively, over an effective area of ≈600 square degrees at 95 GHz and ≈400 square degrees at 150 and 220 GHz. The 220 GHz maps now achieve a signal-to-noise ratio on polarized dust emission exceeding that of Planck at 353 GHz. We take auto- and cross-spectra between these maps and publicly available WMAP and Planck maps at frequencies from 23 to 353 GHz and evaluate the joint likelihood of the spectra versus a multicomponent model of lensed ΛCDM+r+dust+synchrotron+noise. The foreground model has seven parameters, and no longer requires a prior on the frequency spectral index of the dust emission taken from measurements on other regions of the sky. This model is an adequate description of the data at the current noise levels. The likelihood analysis yields the constraint r_{0.05}<0.036 at 95% confidence. Running maximum likelihood search on simulations we obtain unbiased results and find that σ(r)=0.009. These are the strongest constraints to date on primordial gravitational waves.

The FAST-M complex intervention for the detection and management of maternal sepsis in low-resource settings: a multi-site evaluation.

OBJECTIVE: To evaluate whether the implementation of the FAST-M complex intervention was feasible and improved the recognition and management of maternal sepsis in a low-resource setting. DESIGN: A before-and-after design. SETTING: Fifteen government healthcare facilities in Malawi. POPULATION: Women suspected of having maternal sepsis. METHODS: The FAST-M complex intervention consisted of the following components: the FAST-M maternal sepsis treatment bundle and the FAST-M implementation programme. Performance of selected process outcomes was compared between a 2-month baseline phase and 6-month intervention phase with compliance used as a proxy measure of feasibility. MAIN OUTCOME RESULT: Compliance with vital sign recording and use of the FAST-M maternal sepsis bundle. RESULTS: Following implementation of the FAST-M intervention, women were more likely to have a complete set of vital signs taken on admission to the wards (0/163 [0%] versus 169/252 [67.1%], P < 0.001). Recognition of suspected maternal sepsis improved with more cases identified following the intervention (12/106 [11.3%] versus 107/166 [64.5%], P < 0.001). Sepsis management improved, with women more likely to receive all components of the FAST-M treatment bundle within 1 hour of recognition (0/12 [0%] versus 21/107 [19.6%], P = 0.091). In particular, women were more likely to receive antibiotics (3/12 [25.0%] versus 72/107 [67.3%], P = 0.004) within 1 hour of recognition of suspected sepsis. CONCLUSION: Implementation of the FAST-M complex intervention was feasible and led to the improved recognition and management of suspected maternal sepsis in a low-resource setting such as Malawi. TWEETABLE ABSTRACT: Implementation of a sepsis care bundle for low-resources improved recognition & management of maternal sepsis.

Broadband, millimeter-wave antireflection coatings for large-format, cryogenic aluminum oxide optics.

We present two prescriptions for broadband ($ {\sim} 77 - 252\;{\rm GHz} $), millimeter-wave antireflection coatings for cryogenic, sintered polycrystalline aluminum oxide optics: one for large-format (700 mm diameter) planar and plano-convex elements, the other for densely packed arrays of quasi-optical elements-in our case, 5 mm diameter half-spheres (called "lenslets"). The coatings comprise three layers of commercially available, polytetrafluoroethylene-based, dielectric sheet material. The lenslet coating is molded to fit the 150 mm diameter arrays directly, while the large-diameter lenses are coated using a tiled approach. We review the fabrication processes for both prescriptions, then discuss laboratory measurements of their transmittance and reflectance. In addition, we present the inferred refractive indices and loss tangents for the coating materials and the aluminum oxide substrate. We find that at 150 GHz and 300 K the large-format coating sample achieves $ (97 \pm 2)\% $ transmittance, and the lenslet coating sample achieves $ (94 \pm 3)\% $ transmittance.

Development of the FAST-M maternal sepsis bundle for use in low-resource settings: a modified Delphi process.

OBJECTIVE: To develop a sepsis care bundle for the initial management of maternal sepsis in low resource settings. DESIGN: Modified Delphi process. SETTING: Participants from 34 countries. POPULATION: Healthcare practitioners working in low resource settings (n = 143; 34 countries), members of an expert panel (n = 11) and consultation with the World Health Organization Global Maternal and Neonatal Sepsis Initiative technical working group. METHODS: We reviewed the literature to identify all potential interventions and practices around the initial management of sepsis that could be bundled together. A modified Delphi process, using an online questionnaire and in-person meetings, was then undertaken to gain consensus on bundle items. Participants ranked potential bundle items in terms of perceived importance and feasibility, considering their use in both hospitals and health centres. Findings from the healthcare practitioners were then triangulated with those of the experts. MAIN OUTCOME MEASURE: Consensus on bundle items. RESULTS: Consensus was reached after three consultation rounds, with the same items deemed most important and feasible by both the healthcare practitioners and expert panel. Final bundle items selected were: (1) Fluids, (2) Antibiotics, (3) Source identification and control, (4) Transfer (to appropriate higher-level care) and (5) Monitoring (of both mother and neonate as appropriate). The bundle was given the acronym 'FAST-M'. CONCLUSION: A clinically relevant maternal sepsis bundle for low resource settings has been developed by international consensus. TWEETABLE ABSTRACT: A maternal sepsis bundle for low resource settings has been developed by international consensus.

The impact of patient demographics and comorbidities upon burns admitted to Tygerberg Hospital Burns Unit, Western Cape, South Africa.

In South Africa, burns are a major public health problem responsible for significant morbidity and long-term physical disability. This is, in part, due to a significant proportion of the urban population living in poorly constructed, combustible accommodation. The presence of co-morbid diseases such as diabetes and malignancy in patients with burns has been associated with a poorer outcome. The impact of other diseases such as HIV has yet to be defined. A retrospective data collection study analysed the 221 patients admitted to Tygerberg Hospital Burns Unit in 2011 and the first six months of 2013. Using hospital records, patient demographic data was collected alongside burn agent, ICU admission, complications, and patient outcome in terms of length of stay and mortality. The most common burn agent was hot liquid (45.7%). A significant proportion of patients were subject to intentional attacks (34.3%). Shack fires and flame accounted cumulatively for 85% of total inhalational burns, the highest rates of admission to ICU (85.5%), the highest rate of complications, as well as 92.3% of all total fatalities. HIV+ patients had a higher mortality (13.3% vs 5%, p=0.22) and a higher complication rate (46.7% vs 30%, p=0.21). There was no difference in length of stay between the HIV+ and HIV- cohort (12days vs. 15.5 days, p=0.916). Burns are a significant yet preventable cause of mortality and morbidity. The rising number of shack fires, responsible for extensive burns and resultant mortality is concerning and indicates urgent attention and action. HIV complicates the recovery from burn and is responsible for an increased rate of in hospital mortality.

Involvement of double-stranded RNA-activated protein kinase in the synergistic activation of nuclear factor-kappaB by tumor necrosis factor-alpha and gamma-interferon in preneuronal cells.

Tumor necrosis factor-alpha (TNF-alpha) and gamma-interferon (IFN-gamma) cooperate during a variety of biological responses and ultimately synergistically enhance the expression of genes involved in immune and inflammatory responses. Recently, we demonstrated that IFN-gamma can significantly potentiate TNF-alpha-induced nuclear factor (NF)-kappaB nuclear translocation in neuronal derived and endothelial cell lines. The mechanism by which these two cytokines exert their synergistic effect on NF-kappaB involves the de novo degradation of the NF-kappaB inhibitor, IkappaBbeta. The double-stranded RNA-dependent kinase PKR is IFN-inducible and has been implicated in the activation of NF-kappaB; therefore, we examined the possibility that PKR may play a role in the synergistic activation of NF-kappaB during TNF-alpha/IFN-gamma cotreatment. The PKR inhibitor 2-aminopurine (2-AP) inhibited TNF-alpha/IFN-gamma-induced NF-kappaB nuclear translocation in neuronal derived cells but not in endothelial cells. The induced degradation of IkappaBbeta, which is normally observed upon TNF-alpha/IFN-gamma cotreatment, was blocked completely by 2-AP in neuronal derived cells. Also, 2-AP treatment or overexpression of a catalytically inactive PKR inhibited the TNF-alpha/IFN-gamma-induced synergistic activation of kappaB-dependent gene expression. Our results suggest that the signal generated by IFN-gamma during TNF-alpha/IFN-gamma cotreatment may require PKR to elicit enhanced NF-kappaB activity, and this signal may affect the stability of the IkappaBbeta protein.

The NPC derived C15 LMP1 protein confers enhanced activation of NF-kappa B and induction of the EGFR in epithelial cells.

The Epstein-Barr Virus (EBV) LMP1 protein is frequently expressed in nasopharyngeal carcinoma and is essential for the transforming effects of EBV. Analysis of LMP1 genes isolated from tumor biopsies has revealed considerable sequence variation including deletion of amino acids 343-352. Several studies have suggested that this sequence variation could enhance the transforming potential of LMP1. LMP1 has profound effects on cellular gene expression mediated in part through activation of the NF-kappa B transcription factor. In addition, LMP1 engages the TRAF signaling pathway resulting in the induction of EGFR expression. In this study, the LMP1 proteins derived from the laboratory strain B95-8 and the NPC strain C15 were analysed for their ability to activate NF-kappa B and also to induce expression of the EGFR. The data suggest that the C15-LMP1 protein activates NF-kappa B more efficiently and induces higher levels of the EGFR. Analysis of chimeric LMP1 proteins indicates that the amino terminal 181 amino acids of C15-LMP1 confer this increased signaling capability, and that deletion of amino acids 343-352 does not affect these properties. Finally, these data provide evidence that five amino acid changes within the transmembrane domain in the C15-LMP1 protein lead to enhanced NF-kappa B activation and EGFR induction.

Interaction of tumor necrosis factor receptor-associated factor signaling proteins with the latent membrane protein 1 PXQXT motif is essential for induction of epidermal growth factor receptor expression.

The Epstein-Barr virus latent membrane protein 1 (LMP1) oncoprotein causes multiple cellular changes, including induction of epidermal growth factor receptor (EGFR) expression and activation of the NF-kappaB transcription factor. LMP1 and the cellular protein CD40, which also induces EGFR expression, interact with the tumor necrosis factor receptor-associated factor (TRAF) proteins. The LMP1 carboxy-terminal activation region 1 signaling domain interacts specifically with the TRAFs and is essential for EGFR induction through a mechanism independent of NF-kappaB alone. LMP1 and CD40 share a common TRAF binding motif, PXQXT. In this study, the PXQXT motifs in both LMP1 and CD40 were altered and mutant proteins were analyzed for induction of EGFR expression. Replacement of the T residue with A in CD40 completely blocked induction of the EGFR, while the same mutation in LMP1 did not affect EGFR induction. Replacement of both P and Q residues with A's in LMP1 reduced EGFR induction by >75%, while deletion of PXQXT blocked EGFR induction. These results genetically link EGFR induction by LMP1 to the TRAF signaling pathway. Overexpression of TRAF2 potently activates NF-kappaB, although TRAF2 did not induce expression of the EGFR either alone or in combination with TRAF1 and TRAF3. In vivo analyses of the interaction of the TRAFs with LMP1 variants mutated in the PXQXT domain indicate that high-level induction of EGFR expression requires interaction with TRAF1, -2, and -3. However, exogenous expression of TRAF3 decreased EGFR induction mediated by either LMP1 or CD40. These data suggest that TRAF-mediated activation of EGFR expression requires assembly of a complex containing the appropriate stoichiometry of TRAF proteins clustered at the cell membrane with LMP1.

Synergistic activation of NF-kappaB by tumor necrosis factor alpha and gamma interferon via enhanced I kappaB alpha degradation and de novo I kappaBbeta degradation.

Tumor necrosis factor alpha (TNF-alpha) and gamma interferon (IFN-gamma) are required for an effective immune response to bacterial infection and these cytokines synergize in a variety of biological responses, including the induction of cytokine, cell adhesion, and inducible nitrous oxide synthase gene expression. Typically, the synergistic effect on gene expression is due to the independent activation of nuclear factor kappaB (NF-kappaB) by TNF-alpha and of signal transducers and activators of transcription or IFN-regulatory factor 1 by IFNs, allowing these transcription factors to bind their unique promoter sites. However, since activation of NF-kappaB by TNF-alpha is often transient and would not activate long-term kappaB-dependent transcription effectively, we explored the effects of IFN-gamma on TNF-alpha-induced NF-kappaB activity. IFN-gamma, which typically does not activate NF-kappaB, synergistically enhanced TNF-alpha-induced NF-kappaB nuclear translocation via a mechanism that involves the induced degradation of I kappaBbeta and that apparently requires tyrosine kinase activity in preneuronal cells but not in endothelial cells. Correspondingly, cotreatment of cells with TNF-alpha and IFN-gamma leads to persistent activation of NF-kappaB and to potent activation of kappaB-dependent gene expression, which may explain, at least in part, the synergy observed between these cytokines, as well as their involvement in the generation of an effective immune response.

Cross-talk between transcription factors NF-kappa B and C/EBP in the transcriptional regulation of genes.

The study of the acute phase response has attracted substantial interest, not only for its medical implication, but also its provision as an excellent system with which to elucidate the molecular mechanisms involved in the modulation of gene expression. Our previous data suggest that the synergistic induction of the major acute phase reactant serum amyloid A2 (SAA2) expression by interleukin-1 (IL-1) and interleukin-6 (IL-6) is mediated by two families of transcription factors, namely NF-kappa B and C/EBP. To understand the molecular mechanisms of this synergy, we have undertaken a molecular dissection of the factors involved in the formation of the regulatory complex. Electrophoretic mobility shift analysis indicates that NF-kappa B p65 (RelA) and p50, but not p52 or c-Rel, bind specifically to the NF-kappa B site of the SAA2 promoter in response to IL-1 stimulation. In addition, C/EBP beta and C/EBP delta, but not C/EBP alpha, bind specifically to the C/EBP site of SAA2 in response to IL-6 stimulation. Transient co-transfection analysis indicates that co-operative association of NF-kappa B p65 with C/EBP beta and, in particular, with C/EBP delta, results in synergistic transcriptional activation of the SAA2 promoter. When incubated together, NF-kappa B p65 and C/EBP beta form a ternary complex by direct protein/protein interaction. Mutational analysis demonstrates that the C-terminus region of the Rel homology domain (RHD) and the C-terminus of the activation domain of p65 are important for its interaction with C/EBP beta. These results suggest the NF-kappa B and C/EBP may form a new complex of transcription factors that mediates the synergistic induction of SAA2 by IL-1 and IL-6.

Ca2+ signaling in the Chlamydomonas flagellar regeneration system: cellular and molecular responses.

In response to certain extracellular stimuli, Chlamydomonas reinhardtii cells excise their flagella, induce expression of more than 200 different flagellar mRNAs, and assemble a new flagellar pair. Normally, flagellar excision, gene induction and outgrowth are tightly coupled temporally. Our previous studies showed that uncoupling the cellular response of flagellar excision from flagellar outgrowth resulted in submaximal flagellar gene induction, and led us to propose that normal flagellar gene induction is a composite response. The present study extends these observations by measuring flagellar gene induction in Chlamydomonas cells stimulated under conditions where both flagellar excision and flagellar outgrowth are blocked. We find that the flagellar genes are induced in a Ca(2+)-dependent manner in response to stimulation in the absence of flagellar excision and outgrowth. Flagellar gene induction is therefore independent of flagellar excision and outgrowth but sensitive to extracellular Ca2+ levels. Thus, flagellar excision, flagellar outgrowth and flagellar gene induction are three responses to a common stimulus that are related by their requirement for extracellular Ca2+.

The role of NF-kappa B and NF-IL6 transactivating factors in the synergistic activation of human serum amyloid A gene expression by interleukin-1 and interleukin-6.

To understand the mechanisms by which large increases in serum amyloid A (SAA) occur during the acute phase response, human hepatoma cells were transfected with SAA2 gene reporter plasmids and stimulated with combinations of cytokines. Although interleukin-1 (IL-1) and interleukin-6 (IL-6) stimulated transcription from this promoter individually, addition of both mediators produced a response between two and nine times greater than the expected additive response. This synergistic activation was dependent on the integrity of at least two cis-acting sequences in the SAA2 enhancer. The SAA2 NF-kappa B site was required functionally for the response to both IL-1 and IL-6 alone as well as for synergistic activation; however, IL-6 did not directly induce binding of nuclear proteins to the NF-kappa B sequence. A NF-IL6 site was required for full induction by IL-1 and IL-6, and also mediated strong transactivation by recombinant NF-IL6. Furthermore, transfected NF-IL6 synergized strongly with co-transfected NF-kappa B, particularly with RelA (p65). However synergy between IL-1 and IL-6 was only partly reduced by mutation of the NF-IL6 site, indicating further levels of interaction in addition to the NF-kappa B/NF-IL6 cooperativity.

Inositol phospholipid metabolism may trigger flagellar excision in Chlamydomonas reinhardtii.

Chlamydomonas reinhardtii cells shed their flagella in response to environmental stress. Under favorable conditions, flagella are quickly regrown. To learn more about the signals that trigger flagellar excision and regrowth we have investigated inositol phospholipid metabolites, molecules implicated in signal transduction in several other systems. After deflagellation by low pH or mastoparan, a potent activator of G proteins, there was a rapid increase in levels of inositol 1,4,5-trisphosphate measured by use of receptor-binding assays and HPLC. This increase was concomitant with a decrease in levels of phosphatidylinositol 4,5-bisphosphate and was followed by an increase in phosphatidic acid, results consistent with activation of phospholipase C and diacylglycerol kinase. Additional experiments suggest that this activated phospholipase C is not important for flagellar regrowth but plays a role in informing the excision apparatus of the environmental stress. Addition of neomycin (an inhibitor of phospholipase C) before exposure of cells to low pH or mastoparan prevented the increase in inositol 1,4,5-trisphosphate and also prevented deflagellation. Addition of neomycin after deflagellation blocked increases in inositol 1,4,5-trisphosphate that normally followed deflagellation, but did not block flagellar assembly. Furthermore, a flagellar excision-defective mutant, fa-1, did not shed its flagella in response to low pH or mastoparan, yet both of these agents activated phospholipase C in these cells. The results suggest that activation of phospholipase C, possibly via a G protein, is a proximal step in the signal transduction pathway inducing deflagellation in Chlamydomonas.

Uncoupling of Chlamydomonas flagellar gene expression and outgrowth from flagellar excision by manipulation of Ca2+.

Chlamydomonas cells respond to certain environmental stimuli by shedding their flagella. Flagellar loss induces a rapid, transient increase in expression of a specific set of genes encoding flagellar proteins, and assembly of a new flagellar pair. While flagellar gene expression and initiation of flagellar outgrowth are normally tightly coupled to flagellar excision, our results demonstrate that these processes can be uncoupled by manipulating Ca2+ levels or calmodulin activity. In our experiments, wild-type cells were stimulated to excise their flagella using mechanical shearing, and at times after deflagellation, flagellar lengths were measured and flagellar mRNA abundance changes were determined by S1 nuclease protection analysis. When extracellular Ca2+ was lowered by addition of EGTA to cultures before excision, flagellar mRNA abundance changes and flagellar outgrowth were temporally uncoupled from flagellar excision. When extracellular Ca2+ was lowered immediately after excision or when calmodulin activity was inhibited with W-7, flagellar outgrowth was uncoupled from flagellar excision and flagellar mRNA abundance changes. Whenever events in the process of flagellar regeneration were temporally uncoupled, the magnitude of the flagellar mRNA abundance change was reduced. These results suggest that flagellar gene expression may be regulated by multiple signals generated from these events, and implicate Ca2+ as a factor in the mechanisms controlling flagellar regeneration.

Constitutive and NF-kappa B-like proteins in the regulation of the serum amyloid A gene by interleukin 1.

Serum amyloid A (SAA) is a major acute-phase protein whose chronic production by the liver can lead to the fatal disorder of secondary amyloidosis. Control of SAA is mediated by several inflammatory cytokines, including interleukin 1 (IL-1). To study the cis-acting regulatory elements responsible for constitutive and IL-1-induced expression, DNA constructs containing varying lengths of the promoter region from the human SAA2 beta gene 5' to the bacterial reporter gene, chloramphenicol acetyltransferase (CAT), were generated and transfected into human hepatoma cells, HepG2. Both positive and negative regulatory elements were found in the 5' flanking region of the human SAA2 beta gene. The more proximal region contains an IL-1 enhancer sequence GGGACTTTCC (SAA kappa B1; between -82 and -91), the binding site for the ubiquitous transcription factor NF-kappa B. IL-1 induction of the binding of nuclear factor to this sequence is maximal between 5 min and 30 min after incubation with IL-1 and negative in cells incubated for 60 min or longer. Mutation of the SAA kappa B1 sequence to a nonbinding form of NF-kappa B (CTCACTTTCC) abolishes the IL-1 effect. The SAA 5' region also contained an upstream repressor element, shown by transfection experiments. Within this element, a second NF-kappa B binding site (SAA kappa B2; -626 to -635) was found, and mutation of SAA kappa B2 to a non-NF-kappa B-binding form results in an increase in both constitutive + IL-1 stimulated SAA transcription.(ABSTRACT TRUNCATED AT 250 WORDS)

Identification of cis-acting sequences responsible for phorbol ester induction of human serum amyloid A gene expression via a nuclear factor kappaB-like transcription factor.

We have analyzed the 5'-flanking region of one of the genes coding for the human acute-phase protein, serum amyloid A (SAA). We found that SAA mRNA could be increased fivefold in transfected cells by treatment with phorbol 12-myristate 13-acetate (PMA). To analyze this observation further, we placed a 265-base-pair 5' SAA fragment upstream of the reporter chloramphenicol acetyltransferase (CAT) gene and transfected this construct into HeLa cells. PMA treatment of these transient transfectants resulted in increased CAT expression. Nuclear proteins from PMA-treated HeLa cells bound to this DNA fragment, and methylation interference analysis showed that the binding was specific to the sequence GGGACTTTCC (between -82 and -91), a sequence previously described by R. Sen and D. Baltimore (Cell 46:705-716, 1986) as the binding site for the nuclear factor NF kappa B. In a cotransfection competition experiment, we could abolish PMA-induced CAT activity by using cloned human immunodeficiency virus long-terminal-repeat DNA containing the NF kappa B-binding sequence. The same long-terminal-repeat DNA containing mutant NF kappa B-binding sequences (G. Nabel and D. Baltimore, Nature [London] 326:711-713, 1987) did not affect CAT expression, which suggested that binding by an NF kappa B-like factor is required for increased SAA transcription.

Anderson Fabry disease, a close linkage with highly polymorphic DNA markers DXS17, DXS87 and DXS88.

Anderson Fabry disease is an X-linked lysosomal storage disorder caused by alpha-galactosidase A deficiency. Hemizygous males and some heterozygous females develop renal failure and cardiovascular complications in early adult life. We have investigated six large UK families to assess the possible linkage of five polymorphic DNA probes to the Anderson Fabry locus, previously localised to Xq21-24. No recombination was found between Anderson Fabry disease and DXS87, DXS88 and DXS17, which gave lodmax = 6.4, 6.4 and 5.8 respectively at theta = 0.10, (upper confidence limit 0.10). DXS3 gave lodmax 2.9 at theta = 0.10 (upper confidence limit 0.25). DXYS1 was excluded from linkage. The best fit map (DXYS1/DXS3) theta = 0.192 (DXS17/DXS87/DXS88/Anderson Fabry locus) provided no information about the order of loci in parentheses due to the absence of recombinants. The close linkage of DXS17, DXS87 and DXS88, together with alpha-galactosidase A estimation, can be used for antenatal diagnosis and carrier detection until the application of a gene specific probe has been evaluated.

Anderson-Fabry disease--family linkage studies using two polymorphic X-linked DNA probes.

Anderson-Fabry disease is an X-linked lysosomal storage disorder due to alpha-galactosidase A deficiency. In affected males there is a high mortality in early adult life due to renal failure and cardiovascular complications. We describe our preliminary results from genetic linkage studies in five families using two polymorphic DNA probes, DXS17 and DXYS1, mapping to an area on the long arm of the X chromosome between Xq13-22. DXS17 identified a Taql polymorphism closely linked to the disease locus in three families (lodmax Z = 4.23. at a recombination fraction decreases theta = 0.0). Restriction fragment length polymorphisms detected by DXYS1 were not linked.

Effects of delipidated serum and lipoprotein-deficient serum on sterol biosynthesis and efflux in cultured skin fibroblasts - a comparison of the behaviour of cells from a control with those from a heterozygote and homozygote for familial hypercholesterolaemia.

The biosynthesis and efflux of sterols from cells into the medium were investigated in skin fibroblasts from a control, a patient with obligate heterozygous familial hypercholesterolaemia and a patient with the homozygous condition. The behaviour of the cells was studied in two lipid free media (lipoprotein deficient and delipidated serum), with and without the addition of low density lipoproteins (LDL) in order to find experimental conditions which showed maximum differences between the three cell lines. Incorporation of [14C]acetate into sterols in the presence (repression) and absence (induction) of LDL was similar in the normal and heterozygous cells, whereas the homozygous cells showed reduced repression and increased induction. In all three cell lines induction of sterol synthesis was greater with delipidated than lipoprotein deficient serum. The efflux of sterols in both the presence and absence of LDL did not differ between the three cell lines, but it was greater when LDL was added to the medium and when delipidated serum was used. Sterol biosynthesis and efflux from the cells of the heterozygote did not differ significantly from those of the control.