DNA-dependent protein kinase (DNA-PK) plays a critical role in non-homologous end joining (NHEJ),... more DNA-dependent protein kinase (DNA-PK) plays a critical role in non-homologous end joining (NHEJ), the predominant pathway that repairs DNA doublestrand breaks (DSB) in response to ionizing radiation (IR) to govern genome integrity. The interaction of the catalytic subunit of DNA-PK (DNA-PKcs) with the K u70 / K u80 heterodimer on DSBs leads to DNA-PK activation; ho we ver, it is not known if upstream signaling events govern this activation. Here, we reveal a regulatory step go verning DNA-PK activ ation b y SIR T2 deacetylation, which facilitates DNA-PKcs localization to DSBs and interaction with Ku, thereby promoting DSB repair by NHEJ. SIRT2 deacetylase activity governs cellular resistance to DSB-inducing agents and promotes NHEJ. SIRT2 furthermore interacts with and deacetylates DNA-PKcs in response to IR. SIRT2 deacetylase activity facilitates DNA-PKcs interaction with Ku and localization to DSBs and pr omotes DNA-PK activ ation and phosphorylation of do wnstream NHEJ substrates. Moreover, tar geting SIRT2 with AGK2, a SIRT2-specific inhibitor, augments the efficacy of IR in cancer cells and tumors. Our findings define a regulatory step for DNA-PK activ ation b y SIR T2-mediated deacetylation, elucidating a critical upstream signaling event initiating the repair of DSBs by NHEJ. Furthermore, our data suggest that SIRT2 inhibition may be a promising rationale-driven therapeutic strategy for increasing the effectiveness of radiation therapy.
medRxiv (Cold Spring Harbor Laboratory), Feb 2, 2022
Rapid, high throughput, automated detection of SARS-CoV-2 neutralizing antibodies against native-... more Rapid, high throughput, automated detection of SARS-CoV-2 neutralizing antibodies against native-like vaccine and delta variant spike trimers
Steroid receptors (SRs) are a closely related family of ligand-dependent nuclear receptors that m... more Steroid receptors (SRs) are a closely related family of ligand-dependent nuclear receptors that mediate the transcription of genes critical for development, reproduction and immunity. SR dysregulation has been implicated in cancer, inflammatory diseases and metabolic disorders. SRs bind their cognate hormone ligand with exquisite specificity, offering a unique system to study the evolution of molecular recognition. The SR family evolved from an estrogen-sensitive ancestor and diverged to become sensitive to progestagens, corticoids and, most recently, androgens. To understand the structural mechanisms driving the evolution of androgen responsiveness, the ancestral androgen receptor (ancAR1) was crystallized in complex with 5-dihydrotestosterone (DHT) and a fragment of the transcriptional mediator/intermediary factor 2 (Tif2). Crystals diffracted to 2.1 Å resolution and the resulting structure will permit a direct comparison with its progestagen-sensitive ancestor, ancestral steroid receptor 2 (AncSR2).
bioRxiv (Cold Spring Harbor Laboratory), May 17, 2019
As a key regulator of metabolism and inflammation, the orphan nuclear hormone receptor, Liver Rec... more As a key regulator of metabolism and inflammation, the orphan nuclear hormone receptor, Liver Receptor Homolog-1 (LRH-1), has potential as a therapeutic target for diabetes, nonalcoholic fatty liver disease, and inflammatory bowel diseases. Discovery of LRH-1 modulators has been difficult, in part due to the tendency for synthetic compounds to bind unpredictably within the lipophilic binding pocket. Using a structure-guided approach, we exploited a newly-discovered polar interaction to lock agonists in a consistent orientation. This enabled the discovery of the first low nanomolar LRH-1 agonist, one hundred times more potent than the best previous modulator. We elucidate a novel mechanism of action that relies upon specific polar interactions deep in the LRH-1 binding pocket. In an organoid model of inflammatory bowel disease, the new agonist increases expression of LRH-1-conrolled steroidogenic genes and promotes anti-inflammatory gene expression changes. These studies constitute major progress in developing LRH-1 modulators with potential clinical utility.
Acta Crystallographica Section F: Structural Biology Communications, Jan 21, 2014
The 1.90 Å resolution X-ray crystal structure of glycerol dehydrogenase derived from contaminatin... more The 1.90 Å resolution X-ray crystal structure of glycerol dehydrogenase derived from contaminating bacteria present during routine Escherichia coli protein expression is presented. This off-target enzyme showed intrinsic affinity for Ni 2+-Sepharose, migrated at the expected molecular mass for the target protein during gel filtration and was crystallized before it was realised that contamination had occurred. In this study, it is shown that liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) can efficiently identify the protein composition of crystals in a crystallization experiment as part of a structure-determination pipeline for an unknown protein. The highresolution X-ray data enabled sequencing directly from the electron-density maps, allowing the source of contamination to be placed within the Serratia genus. Incorporating additional protein-identity checks, such as tandem LC-MS/ MS, earlier in the protein expression, purification and crystallization workflow may have prevented the unintentional structure determination of this metabolic enzyme, which represents the first enterobacterial glycerol dehydrogenase reported to date.
The following experimental techniques were used to determine the structure: X-RAY DIFFRACTION The... more The following experimental techniques were used to determine the structure: X-RAY DIFFRACTION The reported resolution of this entry is 3.00 Å. Percentile scores (ranging between 0-100) for global validation metrics of the entry are shown in the following graphic. The table shows the number of entries on which the scores are based. Metric Whole archive (#Entries) Similar resolution (#Entries, resolution range(Å)) R f ree
The glucocorticoid receptor (GR) is a steroid hormone-activated transcription factor that binds t... more The glucocorticoid receptor (GR) is a steroid hormone-activated transcription factor that binds to various glucocorticoid response elements to up- or down- regulate the transcription of thousands of genes involved in metabolism, development, stress and inflammatory responses. GR consists of two domains enabling interaction with glucocorticoids, DNA response elements and coregulators, as well as a large intrinsically disordered region that mediates condensate formation. A growing body of structural studies during the past decade have shed new light on GR interactions, providing a new understanding of the mechanisms driving context-specific GR activity. Here, we summarize the established and emerging mechanisms of action of GR, primarily from a structural perspective. This minireview also discusses how the current state of knowledge of GR function may guide future glucocorticoid design with an improved therapeutic index for different inflammatory disorders.
Multiple factors in addition to over consumption lead to obesity and non-alcoholic fatty liver di... more Multiple factors in addition to over consumption lead to obesity and non-alcoholic fatty liver disease (NAFLD) in the United States and worldwide. CYP2B6 is the only human detoxification CYP whose loss is associated with obesity, and Cyp2b-null mice show greater diet-induced obesity with increased steatosis than wildtype mice. However, a putative mechanism has not been determined. LC-MS/MS revealed that CYP2B6 metabolizes PUFAs, with a preference for metabolism of ALA to 9-HOTrE and to a lesser extent 13-HOTrE with a preference for metabolism of PUFAs at the 9- and 13-positions. To further study the role of CYP2B6 in vivo, humanized-CYP2B6-transgenic (hCYP2B6-Tg) and Cyp2b-null mice were fed a 60% high-fat diet for 16 weeks. Compared to Cyp2b-null mice, hCYP2B6-Tg mice showed reduced weight gain and metabolic disease as measured by glucose tolerance tests, however hCYP2B6-Tg male mice showed increased liver triglycerides. Serum and liver oxylipin metabolite concentrations increased ...
In this study, by characterizing several human monoclonal antibodies (mAbs) isolated from single ... more In this study, by characterizing several human monoclonal antibodies (mAbs) isolated from single B cells of the COVID-19–recovered individuals in India who experienced ancestral Wuhan strain (WA.1) of SARS-CoV-2 during early stages of the pandemic, we found a receptor binding domain (RBD)–specific mAb 002-S21F2 that has rare gene usage and potently neutralized live viral isolates of SARS-CoV-2 variants including Alpha, Beta, Gamma, Delta, and Omicron sublineages (BA.1, BA.2, BA.2.12.1, BA.4, and BA.5) with IC 50 ranging from 0.02 to 0.13 μg/ml. Structural studies of 002-S21F2 in complex with spike trimers of Omicron and WA.1 showed that it targets a conformationally conserved epitope on the outer face of RBD (class 3 surface) outside the ACE2-binding motif, thereby providing a mechanistic insights for its broad neutralization activity. The discovery of 002-S21F2 and the broadly neutralizing epitope it targets have timely implications for developing a broad range of therapeutic and v...
Widespread and frequent testing is critical to prevent the spread of COVID-19, and rapid antigen ... more Widespread and frequent testing is critical to prevent the spread of COVID-19, and rapid antigen tests are the diagnostic tool of choice in many settings. With new viral variants continuously emerging and spreading rapidly, the effect of mutations on antigen test performance is a major concern. In response to the spread of variants the National Institutes of Health’s Rapid Acceleration of Diagnostics (RADx®) initiative created a Variant Task Force to assess the impact of emerging SARS-CoV-2 variants on in vitro diagnostic testing. To evaluate the impact of mutations on rapid antigen tests we developed a lentivirus-mediated mammalian surface-display platform for the SARS-CoV-2 Nucleocapsid protein, the target of the majority of rapid antigen tests. We employed deep mutational scanning (DMS) to directly measure the effect of all possible Nucleocapsid point mutations on antibody binding by 17 diagnostic antibodies used in 11 commercially available antigen tests with FDA emergency use a...
The effects of age (9 months old), high-fat diet (4.5 months old), and the loss of Cyp2b9, Cyp2b1... more The effects of age (9 months old), high-fat diet (4.5 months old), and the loss of Cyp2b9, Cyp2b10, and Cyp2b13 (Cyp2b-null mice) on the male murine hepatic lipidome was compared. Hierarchical clustering and principal component analysis show that age perturbs phospholipid profiles and serum lipid markers the most compared to healthy, young mice; followed by a high-fat diet and then loss of Cyp2b. WT and Cyp2b-null male mice were divided into groups and fed either commercially available diets; normal chow diet (ND; 2018S-Envigo Teklad Diet) or a high-fat diet (HFD; Envigo TD.06414) for 10 weeks. Mice were 4.5 months old at the end of the HFD study and referred to as ND-fed young or HFD-fed young; WT or Cyp2b-null mice. An additional experimental group of WT and Cyp2b-null male mice were fed a ND until they reached 9 months (termed old). Serum biomarkers and liver triglycerides (TAG) were measured. Liver and inguinal white adipose tissue (WAT) were excised, weighed, and divided by total body weight to determine the hepatosomatic index (HSI) and white adipose somatic index (WSI). Mass spectrometry performed using a Sciex AC LC system and Sciex QTrap5500 mass spectrometer. Phospholipid species were identified and quantified from the livers of mice (n=3 for young (4.5 mo) ND and HFD-fed mice; n=5 for old from each experimental group by LC-MS/MS at the Emory Integrated Lipidomics Core (EILC) as described in their online protocols. Supplementary File 1. Lipidomic data from all treatment groups. Lipid species were identified from liver samples of male mice from arachidonic (ARA), linoleic (LA), α-linolenic (ALA) and docosahexaenoic acid (DHA). Treatment groups are ND-fed young (4.5 mo), ND-fed old (9 mo), and young mice fed a HFD for 10 weeks. All data is in relative percent. Supplementary File 2. Data used to make Figure 4 principal component analysis (PCA) biplot. Supplementary File 3. (A) Lipids that are statistically different by Student's t-tests between Cyp2b-null and WT mice in the different age and diet gr [...]
BACKGROUND & AIMS A Western-style diet, which is high in fat and sugar, can cause significant... more BACKGROUND & AIMS A Western-style diet, which is high in fat and sugar, can cause significant dyslipidemia and non-alcoholic fatty liver disease; the diet has an especially strong effect in women, regardless of total calorie intake. Dietary supplementation with beneficial microbes might reduce the detrimental effects of a Western-style diet. We assessed the effects of Lactococcus lactis subsp. cremoris on weight gain, liver fat, serum cholesterol, and insulin resistance in female mice on a high-fat, high-carbohydrate diet. METHODS Female C57BL/6 mice were fed either a high-fat, high-carbohydrate (Western-style) diet that contained 40% fat, (mostly milk fat) and 43% carbohydrate (mostly sucrose) or calorie-matched per gram control diet. The diets of mice were supplemented with 1x 109 CFU of L lactis subsp. cremoris ATCC 19257 or Lactobacillus rhamnosus GG ATCC 53103 (control bacteria), 3 times per week for 16 weeks. Body weights were measured, and fecal, blood, and liver tissues were collected and analyzed. Livers were analyzed for fat accumulation and inflammation and blood samples were analyzed for cholesterol and glucose levels. Mice were housed within CLAMS-HC monitoring systems and respiratory exchange ratio and activity measured. Hepatic lipid profiles of L lactis subsp. cremoris-supplemented mice were characterized by lipidomic mass spectrometry analysis. RESULTS Mice fed L lactis subsp. cremoris while on the Western-style diet gained less weight, developed less hepatic steatosis and inflammation, and had a lower mean serum level of cholesterol and body mass index than mice fed the control bacteria. Mice fed the L lactis subsp. cremoris had increased glucose intolerance while on the Western-style diet, compared to mice fed control bacteria, and had alterations in hepatic lipids, including oxylipins. CONCLUSIONS Dietary supplementation with L lactis subsp. cremoris in female mice on a high-fat, high-carbohydrate (Western-style) diet caused them to gain less weight, develop less liver fat an inflammation, reduce serum cholesterol levels, and increase glucose tolerance, compared with mice on the same diet fed control bacteria. L lactis subsp. cremoris is safe for oral ingestion and might be developed for persons with metabolic and liver disorders caused by a Western-style diet.
DNA-dependent protein kinase (DNA-PK) plays a critical role in non-homologous end joining (NHEJ),... more DNA-dependent protein kinase (DNA-PK) plays a critical role in non-homologous end joining (NHEJ), the predominant pathway that repairs DNA doublestrand breaks (DSB) in response to ionizing radiation (IR) to govern genome integrity. The interaction of the catalytic subunit of DNA-PK (DNA-PKcs) with the K u70 / K u80 heterodimer on DSBs leads to DNA-PK activation; ho we ver, it is not known if upstream signaling events govern this activation. Here, we reveal a regulatory step go verning DNA-PK activ ation b y SIR T2 deacetylation, which facilitates DNA-PKcs localization to DSBs and interaction with Ku, thereby promoting DSB repair by NHEJ. SIRT2 deacetylase activity governs cellular resistance to DSB-inducing agents and promotes NHEJ. SIRT2 furthermore interacts with and deacetylates DNA-PKcs in response to IR. SIRT2 deacetylase activity facilitates DNA-PKcs interaction with Ku and localization to DSBs and pr omotes DNA-PK activ ation and phosphorylation of do wnstream NHEJ substrates. Moreover, tar geting SIRT2 with AGK2, a SIRT2-specific inhibitor, augments the efficacy of IR in cancer cells and tumors. Our findings define a regulatory step for DNA-PK activ ation b y SIR T2-mediated deacetylation, elucidating a critical upstream signaling event initiating the repair of DSBs by NHEJ. Furthermore, our data suggest that SIRT2 inhibition may be a promising rationale-driven therapeutic strategy for increasing the effectiveness of radiation therapy.
medRxiv (Cold Spring Harbor Laboratory), Feb 2, 2022
Rapid, high throughput, automated detection of SARS-CoV-2 neutralizing antibodies against native-... more Rapid, high throughput, automated detection of SARS-CoV-2 neutralizing antibodies against native-like vaccine and delta variant spike trimers
Steroid receptors (SRs) are a closely related family of ligand-dependent nuclear receptors that m... more Steroid receptors (SRs) are a closely related family of ligand-dependent nuclear receptors that mediate the transcription of genes critical for development, reproduction and immunity. SR dysregulation has been implicated in cancer, inflammatory diseases and metabolic disorders. SRs bind their cognate hormone ligand with exquisite specificity, offering a unique system to study the evolution of molecular recognition. The SR family evolved from an estrogen-sensitive ancestor and diverged to become sensitive to progestagens, corticoids and, most recently, androgens. To understand the structural mechanisms driving the evolution of androgen responsiveness, the ancestral androgen receptor (ancAR1) was crystallized in complex with 5-dihydrotestosterone (DHT) and a fragment of the transcriptional mediator/intermediary factor 2 (Tif2). Crystals diffracted to 2.1 Å resolution and the resulting structure will permit a direct comparison with its progestagen-sensitive ancestor, ancestral steroid receptor 2 (AncSR2).
bioRxiv (Cold Spring Harbor Laboratory), May 17, 2019
As a key regulator of metabolism and inflammation, the orphan nuclear hormone receptor, Liver Rec... more As a key regulator of metabolism and inflammation, the orphan nuclear hormone receptor, Liver Receptor Homolog-1 (LRH-1), has potential as a therapeutic target for diabetes, nonalcoholic fatty liver disease, and inflammatory bowel diseases. Discovery of LRH-1 modulators has been difficult, in part due to the tendency for synthetic compounds to bind unpredictably within the lipophilic binding pocket. Using a structure-guided approach, we exploited a newly-discovered polar interaction to lock agonists in a consistent orientation. This enabled the discovery of the first low nanomolar LRH-1 agonist, one hundred times more potent than the best previous modulator. We elucidate a novel mechanism of action that relies upon specific polar interactions deep in the LRH-1 binding pocket. In an organoid model of inflammatory bowel disease, the new agonist increases expression of LRH-1-conrolled steroidogenic genes and promotes anti-inflammatory gene expression changes. These studies constitute major progress in developing LRH-1 modulators with potential clinical utility.
Acta Crystallographica Section F: Structural Biology Communications, Jan 21, 2014
The 1.90 Å resolution X-ray crystal structure of glycerol dehydrogenase derived from contaminatin... more The 1.90 Å resolution X-ray crystal structure of glycerol dehydrogenase derived from contaminating bacteria present during routine Escherichia coli protein expression is presented. This off-target enzyme showed intrinsic affinity for Ni 2+-Sepharose, migrated at the expected molecular mass for the target protein during gel filtration and was crystallized before it was realised that contamination had occurred. In this study, it is shown that liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) can efficiently identify the protein composition of crystals in a crystallization experiment as part of a structure-determination pipeline for an unknown protein. The highresolution X-ray data enabled sequencing directly from the electron-density maps, allowing the source of contamination to be placed within the Serratia genus. Incorporating additional protein-identity checks, such as tandem LC-MS/ MS, earlier in the protein expression, purification and crystallization workflow may have prevented the unintentional structure determination of this metabolic enzyme, which represents the first enterobacterial glycerol dehydrogenase reported to date.
The following experimental techniques were used to determine the structure: X-RAY DIFFRACTION The... more The following experimental techniques were used to determine the structure: X-RAY DIFFRACTION The reported resolution of this entry is 3.00 Å. Percentile scores (ranging between 0-100) for global validation metrics of the entry are shown in the following graphic. The table shows the number of entries on which the scores are based. Metric Whole archive (#Entries) Similar resolution (#Entries, resolution range(Å)) R f ree
The glucocorticoid receptor (GR) is a steroid hormone-activated transcription factor that binds t... more The glucocorticoid receptor (GR) is a steroid hormone-activated transcription factor that binds to various glucocorticoid response elements to up- or down- regulate the transcription of thousands of genes involved in metabolism, development, stress and inflammatory responses. GR consists of two domains enabling interaction with glucocorticoids, DNA response elements and coregulators, as well as a large intrinsically disordered region that mediates condensate formation. A growing body of structural studies during the past decade have shed new light on GR interactions, providing a new understanding of the mechanisms driving context-specific GR activity. Here, we summarize the established and emerging mechanisms of action of GR, primarily from a structural perspective. This minireview also discusses how the current state of knowledge of GR function may guide future glucocorticoid design with an improved therapeutic index for different inflammatory disorders.
Multiple factors in addition to over consumption lead to obesity and non-alcoholic fatty liver di... more Multiple factors in addition to over consumption lead to obesity and non-alcoholic fatty liver disease (NAFLD) in the United States and worldwide. CYP2B6 is the only human detoxification CYP whose loss is associated with obesity, and Cyp2b-null mice show greater diet-induced obesity with increased steatosis than wildtype mice. However, a putative mechanism has not been determined. LC-MS/MS revealed that CYP2B6 metabolizes PUFAs, with a preference for metabolism of ALA to 9-HOTrE and to a lesser extent 13-HOTrE with a preference for metabolism of PUFAs at the 9- and 13-positions. To further study the role of CYP2B6 in vivo, humanized-CYP2B6-transgenic (hCYP2B6-Tg) and Cyp2b-null mice were fed a 60% high-fat diet for 16 weeks. Compared to Cyp2b-null mice, hCYP2B6-Tg mice showed reduced weight gain and metabolic disease as measured by glucose tolerance tests, however hCYP2B6-Tg male mice showed increased liver triglycerides. Serum and liver oxylipin metabolite concentrations increased ...
In this study, by characterizing several human monoclonal antibodies (mAbs) isolated from single ... more In this study, by characterizing several human monoclonal antibodies (mAbs) isolated from single B cells of the COVID-19–recovered individuals in India who experienced ancestral Wuhan strain (WA.1) of SARS-CoV-2 during early stages of the pandemic, we found a receptor binding domain (RBD)–specific mAb 002-S21F2 that has rare gene usage and potently neutralized live viral isolates of SARS-CoV-2 variants including Alpha, Beta, Gamma, Delta, and Omicron sublineages (BA.1, BA.2, BA.2.12.1, BA.4, and BA.5) with IC 50 ranging from 0.02 to 0.13 μg/ml. Structural studies of 002-S21F2 in complex with spike trimers of Omicron and WA.1 showed that it targets a conformationally conserved epitope on the outer face of RBD (class 3 surface) outside the ACE2-binding motif, thereby providing a mechanistic insights for its broad neutralization activity. The discovery of 002-S21F2 and the broadly neutralizing epitope it targets have timely implications for developing a broad range of therapeutic and v...
Widespread and frequent testing is critical to prevent the spread of COVID-19, and rapid antigen ... more Widespread and frequent testing is critical to prevent the spread of COVID-19, and rapid antigen tests are the diagnostic tool of choice in many settings. With new viral variants continuously emerging and spreading rapidly, the effect of mutations on antigen test performance is a major concern. In response to the spread of variants the National Institutes of Health’s Rapid Acceleration of Diagnostics (RADx®) initiative created a Variant Task Force to assess the impact of emerging SARS-CoV-2 variants on in vitro diagnostic testing. To evaluate the impact of mutations on rapid antigen tests we developed a lentivirus-mediated mammalian surface-display platform for the SARS-CoV-2 Nucleocapsid protein, the target of the majority of rapid antigen tests. We employed deep mutational scanning (DMS) to directly measure the effect of all possible Nucleocapsid point mutations on antibody binding by 17 diagnostic antibodies used in 11 commercially available antigen tests with FDA emergency use a...
The effects of age (9 months old), high-fat diet (4.5 months old), and the loss of Cyp2b9, Cyp2b1... more The effects of age (9 months old), high-fat diet (4.5 months old), and the loss of Cyp2b9, Cyp2b10, and Cyp2b13 (Cyp2b-null mice) on the male murine hepatic lipidome was compared. Hierarchical clustering and principal component analysis show that age perturbs phospholipid profiles and serum lipid markers the most compared to healthy, young mice; followed by a high-fat diet and then loss of Cyp2b. WT and Cyp2b-null male mice were divided into groups and fed either commercially available diets; normal chow diet (ND; 2018S-Envigo Teklad Diet) or a high-fat diet (HFD; Envigo TD.06414) for 10 weeks. Mice were 4.5 months old at the end of the HFD study and referred to as ND-fed young or HFD-fed young; WT or Cyp2b-null mice. An additional experimental group of WT and Cyp2b-null male mice were fed a ND until they reached 9 months (termed old). Serum biomarkers and liver triglycerides (TAG) were measured. Liver and inguinal white adipose tissue (WAT) were excised, weighed, and divided by total body weight to determine the hepatosomatic index (HSI) and white adipose somatic index (WSI). Mass spectrometry performed using a Sciex AC LC system and Sciex QTrap5500 mass spectrometer. Phospholipid species were identified and quantified from the livers of mice (n=3 for young (4.5 mo) ND and HFD-fed mice; n=5 for old from each experimental group by LC-MS/MS at the Emory Integrated Lipidomics Core (EILC) as described in their online protocols. Supplementary File 1. Lipidomic data from all treatment groups. Lipid species were identified from liver samples of male mice from arachidonic (ARA), linoleic (LA), α-linolenic (ALA) and docosahexaenoic acid (DHA). Treatment groups are ND-fed young (4.5 mo), ND-fed old (9 mo), and young mice fed a HFD for 10 weeks. All data is in relative percent. Supplementary File 2. Data used to make Figure 4 principal component analysis (PCA) biplot. Supplementary File 3. (A) Lipids that are statistically different by Student's t-tests between Cyp2b-null and WT mice in the different age and diet gr [...]
BACKGROUND & AIMS A Western-style diet, which is high in fat and sugar, can cause significant... more BACKGROUND & AIMS A Western-style diet, which is high in fat and sugar, can cause significant dyslipidemia and non-alcoholic fatty liver disease; the diet has an especially strong effect in women, regardless of total calorie intake. Dietary supplementation with beneficial microbes might reduce the detrimental effects of a Western-style diet. We assessed the effects of Lactococcus lactis subsp. cremoris on weight gain, liver fat, serum cholesterol, and insulin resistance in female mice on a high-fat, high-carbohydrate diet. METHODS Female C57BL/6 mice were fed either a high-fat, high-carbohydrate (Western-style) diet that contained 40% fat, (mostly milk fat) and 43% carbohydrate (mostly sucrose) or calorie-matched per gram control diet. The diets of mice were supplemented with 1x 109 CFU of L lactis subsp. cremoris ATCC 19257 or Lactobacillus rhamnosus GG ATCC 53103 (control bacteria), 3 times per week for 16 weeks. Body weights were measured, and fecal, blood, and liver tissues were collected and analyzed. Livers were analyzed for fat accumulation and inflammation and blood samples were analyzed for cholesterol and glucose levels. Mice were housed within CLAMS-HC monitoring systems and respiratory exchange ratio and activity measured. Hepatic lipid profiles of L lactis subsp. cremoris-supplemented mice were characterized by lipidomic mass spectrometry analysis. RESULTS Mice fed L lactis subsp. cremoris while on the Western-style diet gained less weight, developed less hepatic steatosis and inflammation, and had a lower mean serum level of cholesterol and body mass index than mice fed the control bacteria. Mice fed the L lactis subsp. cremoris had increased glucose intolerance while on the Western-style diet, compared to mice fed control bacteria, and had alterations in hepatic lipids, including oxylipins. CONCLUSIONS Dietary supplementation with L lactis subsp. cremoris in female mice on a high-fat, high-carbohydrate (Western-style) diet caused them to gain less weight, develop less liver fat an inflammation, reduce serum cholesterol levels, and increase glucose tolerance, compared with mice on the same diet fed control bacteria. L lactis subsp. cremoris is safe for oral ingestion and might be developed for persons with metabolic and liver disorders caused by a Western-style diet.
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Papers by Eric Ortlund