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Patrick P. Lowe, Benedek Gyongyosi, Abhishek Satishchandran, Arvin Iracheta-Vellve, Aditya Ambade, Karen Kodys, Donna Catalano, Doyle V. Ward, Gyongyi Szabo
Alcohol-related changes in the intestinal microbiome influence neutrophil infiltration, inflammation and steatosis in early alcoholic hepatitis in mice
www.plosone.org
www.plosone.org
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(of)Tj
1.0148 0 Td
(liquid)Tj
2.5625 0 Td
(diet)Tj
1.7348 0 Td
(and)Tj
-35.1097 -1.3039 Td
(continuing)Tj
4.6204 0 Td
(until)Tj
2.109 0 Td
(the)Tj
1.4683 0 Td
(oral)Tj
1.7801 0 Td
(ethanol)Tj
3.2032 0 Td
(or)Tj
1.0998 0 Td
(maltose)Tj
3.3109 0 Td
(dextrin)Tj
3.118 0 Td
(gavage)Tj
2.897 0 Td
(on)Tj
1.2756 0 Td
(the)Tj
1.4684 0 Td
(final)Tj
2.0069 0 Td
(feeding)Tj
3.1634 0 Td
(day.)Tj
1.8595 0 Td
(Non-anti-)Tj
-33.3805 -1.2982 Td
(biotic-treated)Tj
5.6182 0 Td
(mice)Tj
2.1486 0 Td
(received)Tj
3.5376 0 Td
(an)Tj
1.2019 0 Td
(equivalent)Tj
4.354 0 Td
(volume)Tj
3.1975 0 Td
(of)Tj
1.0148 0 Td
(water.)Tj
2.6532 0 Td
(Gavage)Tj
3.1464 0 Td
(needles)Tj
3.1748 0 Td
(were)Tj
2.126 0 Td
(rinsed)Tj
2.7155 0 Td
(with)Tj
-34.8885 -1.2983 Td
(acid-treated)Tj
4.9889 0 Td
(water)Tj
2.4264 0 Td
(and)Tj
1.7235 0 Td
(sterilized)Tj
3.8097 0 Td
(in)Tj
1.0318 0 Td
(a)Tj
0.6463 0 Td
(hot-bead)Tj
3.7871 0 Td
(sterilizer)Tj
3.6567 0 Td
(between)Tj
3.5206 0 Td
(each)Tj
2.0182 0 Td
(mouse)Tj
2.8517 0 Td
(treatment)Tj
4.1272 0 Td
(to)Tj
1.0204 0 Td
(min-)Tj
-35.6085 -1.2983 Td
(imize)Tj
2.4094 0 Td
(transfer)Tj
3.3165 0 Td
(of)Tj
1.0148 0 Td
(microbes)Tj
3.9004 0 Td
(between)Tj
3.5263 0 Td
(animals.)Tj
-12.9712 -1.3039 Td
(Feces)Tj
2.3811 0 Td
(were)Tj
2.1203 0 Td
(collected)Tj
3.7417 0 Td
(directly)Tj
3.2314 0 Td
(from)Tj
2.2054 0 Td
(the)Tj
1.474 0 Td
(anus)Tj
2.0863 0 Td
(prior)Tj
2.2563 0 Td
(to)Tj
1.0262 0 Td
(euthanization)Tj
5.7146 0 Td
(and)Tj
1.7291 0 Td
(frozen)Tj
2.7723 0 Td
(at)Tj
0.9524 0 Td
(-20C)Tj
2.5795 0 Td
(until)Tj
-35.4668 -1.2982 Td
(plating.)Tj
3.2201 0 Td
(Upon)Tj
2.5285 0 Td
(thawing,)Tj
3.6737 0 Td
(stool)Tj
2.1373 0 Td
(was)Tj
1.7007 0 Td
(immediately)Tj
5.1987 0 Td
(weighed,)Tj
3.7644 0 Td
(dissociated)Tj
4.6205 0 Td
(in)Tj
1.0318 0 Td
(liquid)Tj
2.5625 0 Td
(thioglycolate)Tj
-30.4382 -1.2983 Td
(media)Tj
2.6928 0 Td
(\(Sigma\))Tj
3.3619 0 Td
(and)Tj
1.7291 0 Td
(diluted)Tj
3.0388 0 Td
(1:1000)Tj
2.8232 0 Td
(prior)Tj
2.2507 0 Td
(to)Tj
1.0262 0 Td
(plating)Tj
2.999 0 Td
(on)Tj
1.2756 0 Td
(non-selective)Tj
5.4935 0 Td
(agar)Tj
1.9276 0 Td
(plates)Tj
2.5001 0 Td
(\(EMD)Tj
2.7553 0 Td
(Milli-)Tj
-33.8738 -1.3039 Td
(pore\).)Tj
2.6021 0 Td
(Plates)Tj
2.5455 0 Td
(were)Tj
2.1203 0 Td
(incubated)Tj
4.1783 0 Td
(at)Tj
0.9524 0 Td
(37C)Tj
2.228 0 Td
(for)Tj
1.3833 0 Td
(48h.)Tj
1.9219 0 Td
(Colonies)Tj
3.7474 0 Td
(were)Tj
2.1203 0 Td
(quantified)Tj
4.3086 0 Td
(using)Tj
2.3924 0 Td
(OpenCFU)Tj
4.3824 0 Td
([)Tj
0.83 0.64 0.02 0 k
(17)Tj
0 g
(])Tj
-34.8829 -1.2983 Td
(and)Tj
1.7291 0 Td
(were)Tj
2.1203 0 Td
(normalized)Tj
4.7792 0 Td
(to)Tj
1.0261 0 Td
(the)Tj
1.4683 0 Td
(mass)Tj
2.1997 0 Td
(of)Tj
1.0148 0 Td
(stool.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 560.0692 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(Histology)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 543.0614 cm
BT
/F8 1 Tf
1 TL
-0.004 Tc
0 0 Td
(OilRed)Tj
3.05 0 Td
(O)Tj
0.9695 0 Td
(tissue)Tj
2.4547 0 Td
(staining)Tj
3.4129 0 Td
(on)Tj
1.2756 0 Td
(OCT-embedded)Tj
6.7691 0 Td
(frozen)Tj
2.7723 0 Td
(liver)Tj
1.9842 0 Td
(sections)Tj
3.4072 0 Td
(was)Tj
1.7008 0 Td
(completed)Tj
4.405 0 Td
(and)Tj
1.7348 0 Td
(quantifi-)Tj
-33.9361 -1.3039 Td
(cation)Tj
2.6985 0 Td
(was)Tj
1.6951 0 Td
(performed)Tj
4.4731 0 Td
(using)Tj
2.3867 0 Td
(ImageJ)Tj
3.0331 0 Td
(to)Tj
1.0261 0 Td
(assess)Tj
2.5228 0 Td
(hepatic)Tj
3.1068 0 Td
(steatosis.)Tj
3.7587 0 Td
(Formalin-fixed)Tj
6.2645 0 Td
(paraffin-embed-)Tj
-30.9654 -1.2982 Td
(ded)Tj
1.6951 0 Td
(liver)Tj
1.9842 0 Td
(sections)Tj
3.4129 0 Td
(were)Tj
2.1203 0 Td
(stained)Tj
3.0784 0 Td
(with)Tj
2.0012 0 Td
(anti-mouse)Tj
4.7565 0 Td
(myeloperoxidase)Tj
6.9676 0 Td
(antibody)Tj
3.7643 0 Td
(\(Abcam\))Tj
3.7701 0 Td
(and)Tj
-33.5506 -1.2983 Td
(subsequently)Tj
5.4028 0 Td
(labeled)Tj
3.0273 0 Td
(with)Tj
2.0013 0 Td
(streptavidin-biotin)Tj
7.7328 0 Td
(immunoenzymatic)Tj
7.7953 0 Td
(antigen)Tj
3.2031 0 Td
(for)Tj
1.3833 0 Td
(detection)Tj
3.9401 0 Td
(with)Tj
-34.486 -1.2983 Td
(3,3'-diaminobenzidine)Tj
9.2578 0 Td
(\(DAB\))Tj
2.9027 0 Td
(\(UltraVision)Tj
5.2894 0 Td
(Mouse)Tj
2.931 0 Td
(Tissue)Tj
2.7723 0 Td
(Detection)Tj
4.1442 0 Td
(System)Tj
3.0557 0 Td
(Anti-Mouse)Tj
-30.3531 -1.3039 Td
(HRP/DAB;)Tj
4.7224 0 Td
(Lab)Tj
1.7008 0 Td
(Vision\).)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 447.7039 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(Biochemical)Tj
5.5417 0 Td
(assays)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 430.696 cm
BT
/F8 1 Tf
1 TL
-0.004 Tc
0 0 Td
(Serum)Tj
2.8346 0 Td
(was)Tj
1.7008 0 Td
(separated)Tj
4.0081 0 Td
(from)Tj
2.211 0 Td
(whole)Tj
2.6079 0 Td
(blood)Tj
2.5115 0 Td
(and)Tj
1.7291 0 Td
(frozen)Tj
2.7723 0 Td
(at)Tj
0.9524 0 Td
(-80C)Tj
2.5795 0 Td
(until)Tj
2.109 0 Td
(use.)Tj
1.7574 0 Td
(Serum)Tj
2.8347 0 Td
(alanine)Tj
3.1237 0 Td
(and)Tj
-33.732 -1.2982 Td
(aspartate)Tj
3.7927 0 Td
(aminotransferases)Tj
7.4664 0 Td
(were)Tj
2.126 0 Td
(measured)Tj
4.1102 0 Td
(using)Tj
2.3867 0 Td
(a)Tj
0.652 0 Td
(kinetic)Tj
2.948 0 Td
(method)Tj
3.3222 0 Td
([)Tj
0.83 0.64 0.02 0 k
(18)Tj
0 g
(].)Tj
2.0749 0 Td
(Serum)Tj
2.8403 0 Td
(endotoxin)Tj
4.3143 0 Td
(was)Tj
-36.0337 -1.3039 Td
(quantified)Tj
4.3086 0 Td
(using)Tj
2.3867 0 Td
(Pierce)Tj
2.6873 0 Td
(LAL)Tj
1.9842 0 Td
(Chromogenic)Tj
5.743 0 Td
(assay)Tj
2.262 0 Td
(\(ThermoFisher\).)Tj
6.8655 0 Td
(Serum)Tj
2.8403 0 Td
(MCP-1)Tj
3.1577 0 Td
(level)Tj
2.0183 0 Td
(was)Tj
-34.2536 -1.2983 Td
(determined)Tj
4.8472 0 Td
(by)Tj
1.1848 0 Td
(ELISA)Tj
2.829 0 Td
(\(BioLegend\).)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 361.3606 cm
BT
/F0 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(mRNA)Tj
3.1417 0 Td
(analysis)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 344.3527 cm
BT
/F8 1 Tf
1 TL
-0.004 Tc
0 0 Td
(RNA)Tj
2.262 0 Td
(was)Tj
1.6951 0 Td
(extracted)Tj
3.8891 0 Td
(from)Tj
2.2053 0 Td
(liver)Tj
1.9843 0 Td
(tissue)Tj
2.4548 0 Td
(using)Tj
2.3924 0 Td
(RNeasy)Tj
3.2485 0 Td
(\(Qiagen\))Tj
3.787 0 Td
(according)Tj
4.1896 0 Td
(to)Tj
1.0205 0 Td
(the)Tj
1.474 0 Td
(manufacturer's)Tj
-30.6026 -1.2982 Td
(instructions,)Tj
5.2213 0 Td
(including)Tj
4.0422 0 Td
(on-column)Tj
4.6998 0 Td
(DNase)Tj
2.914 0 Td
(digestion)Tj
3.8835 0 Td
(\(Zymo)Tj
2.9423 0 Td
(Research\).)Tj
4.371 0 Td
(cDNA)Tj
2.8006 0 Td
(was)Tj
1.7008 0 Td
(written)Tj
-32.5755 -1.3039 Td
(from)Tj
2.2053 0 Td
(1)Tj
/F15 1 Tf
[()]TJ
/F8 1 Tf
(g)Tj
1.6668 0 Td
(of)Tj
1.0147 0 Td
(RNA)Tj
2.2621 0 Td
(and)Tj
1.7291 0 Td
(then)Tj
2.0183 0 Td
(diluted)Tj
3.0387 0 Td
(1:5)Tj
1.4003 0 Td
(in)Tj
1.0318 0 Td
(nuclease-free)Tj
5.4652 0 Td
(water.)Tj
2.6588 0 Td
(SYBR)Tj
2.5285 0 Td
(Green)Tj
2.7043 0 Td
(\(BioRad\))Tj
3.8324 0 Td
(real-time)Tj
-33.5563 -1.2983 Td
(qPCR)Tj
2.5625 0 Td
(was)Tj
1.6951 0 Td
(performed)Tj
4.4617 0 Td
(according)Tj
4.1782 0 Td
(to)Tj
1.0205 0 Td
(the)Tj
1.4626 0 Td
(manufacturer's)Tj
6.2589 0 Td
(instructions.)Tj
5.2157 0 Td
(The)Tj
1.7745 0 Td
(primers)Tj
3.3449 0 Td
(used)Tj
2.0522 0 Td
(are)Tj
1.4457 0 Td
(listed)Tj
-35.4725 -1.2983 Td
(in)Tj
0.83 0.64 0.02 0 k
1.0318 0 Td
(Table)Tj
2.4434 0 Td
(1)Tj
0 g
0.6973 0 Td
(and)Tj
1.7291 0 Td
(18S)Tj
1.6498 0 Td
(was)Tj
1.6951 0 Td
(used)Tj
2.0636 0 Td
(as)Tj
1.0148 0 Td
(a)Tj
0.652 0 Td
(housekeeping)Tj
5.7089 0 Td
(gene)Tj
2.0806 0 Td
(for)Tj
/F13 1 Tf
1.3833 0 Td
[( )]TJ
ET
Q
q
1 j
1 J
0 w
7.4999 0 0 6.6659 426.1606 296.7874 cm
BT
/F13 1 Tf
1 TL
-0.0053 Tc
0 0 Td
[()4()]TJ
ET
Q
q
1 j
1 J
0 w
10 0 0 10 445.1527 292.3653 cm
BT
/F8 1 Tf
1 TL
-0.004 Tc
0 0 Td
(method)Tj
3.3221 0 Td
(of)Tj
1.0148 0 Td
(RNA)Tj
2.2621 0 Td
(expression)Tj
-31.1128 -1.3039 Td
(analysis.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 248.9952 cm
BT
/F2 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(Results)Tj
/F0 1 Tf
0 -1.4976 Td
(Alpha-)Tj
3.0708 0 Td
(and)Tj
1.8662 0 Td
(beta-diversity)Tj
6.0945 0 Td
(of)Tj
1.0441 0 Td
(cecal)Tj
2.5276 0 Td
(microbiome)Tj
5.3245 0 Td
(are)Tj
1.6488 0 Td
(unaltered)Tj
4.3323 0 Td
(by)Tj
1.2662 0 Td
(acute-)Tj
-27.175 -1.1669 Td
(on-chronic)Tj
4.8283 0 Td
(alcohol)Tj
3.3402 0 Td
(feeding)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 200.0125 cm
BT
/F8 1 Tf
1 TL
-0.004 Tc
0 0 Td
(We)Tj
1.61 0 Td
(applied)Tj
3.1521 0 Td
(16S)Tj
1.6441 0 Td
(rDNA)Tj
2.7496 0 Td
(sequencing)Tj
4.7168 0 Td
(to)Tj
1.0262 0 Td
(define)Tj
2.6985 0 Td
(and)Tj
1.7291 0 Td
(to)Tj
1.0262 0 Td
(assess)Tj
2.5171 0 Td
(the)Tj
1.474 0 Td
(changes)Tj
3.4016 0 Td
(in)Tj
1.0375 0 Td
(the)Tj
1.4683 0 Td
(cecal)Tj
2.1713 0 Td
(microbiota)Tj
-32.4224 -1.3039 Td
(induced)Tj
3.4639 0 Td
(by)Tj
1.1848 0 Td
(the)Tj
1.4684 0 Td
(acute-on-chronic)Tj
7.1489 0 Td
(alcohol)Tj
3.1124 0 Td
(feeding)Tj
3.1635 0 Td
(in)Tj
1.0374 0 Td
(mice.)Tj
2.3811 0 Td
(The)Tj
1.7802 0 Td
(bacterial)Tj
3.6226 0 Td
(load)Tj
1.9389 0 Td
(was)Tj
1.6951 0 Td
(substantially)Tj
-31.9972 -1.2982 Td
(reduced)Tj
3.4412 0 Td
(in)Tj
1.0375 0 Td
(antibiotic-treated)Tj
7.1659 0 Td
(mice)Tj
2.1543 0 Td
(and)Tj
1.7291 0 Td
(16S)Tj
1.6498 0 Td
(rDNA)Tj
2.7439 0 Td
(sequence)Tj
3.8608 0 Td
(was)Tj
1.7008 0 Td
(not)Tj
1.5703 0 Td
(obtained)Tj
3.7361 0 Td
(for)Tj
1.3833 0 Td
(that)Tj
1.7801 0 Td
(cohort.)Tj
-33.9531 -1.2983 Td
(Of)Tj
1.2529 0 Td
(the)Tj
1.4683 0 Td
(non-antibiotic-treated)Tj
9.1218 0 Td
(mice,)Tj
2.3811 0 Td
(samples)Tj
3.3789 0 Td
(from)Tj
2.2053 0 Td
(7/8)Tj
1.5024 0 Td
(pair-fed)Tj
3.4015 0 Td
(and)Tj
1.7292 0 Td
(9/10)Tj
1.9785 0 Td
(alcohol-fed)Tj
4.6998 0 Td
(mice)Tj
2.1544 0 Td
(were)Tj
-35.2741 -1.2983 Td
(available)Tj
3.668 0 Td
(for)Tj
1.3833 0 Td
(analysis.)Tj
3.5546 0 Td
(We)Tj
1.6044 0 Td
(observed)Tj
3.7984 0 Td
(no)Tj
1.2756 0 Td
(significant)Tj
4.388 0 Td
(differences)Tj
4.5637 0 Td
(\(p)Tj
/F16 1 Tf
0 Tc
(>)Tj
/F8 1 Tf
-0.004 Tc
(0.05\))Tj
3.8438 0 Td
(in)Tj
/F14 1 Tf
1.0318 0 Td
()Tj
/F8 1 Tf
(-diversity)Tj
4.5694 0 Td
(between)Tj
-33.681 -1.3039 Td
(pair-fed)Tj
3.3958 0 Td
(and)Tj
1.7348 0 Td
(alcohol-fed)Tj
4.6998 0 Td
(groups)Tj
2.9707 0 Td
(\()Tj
0.83 0.64 0.02 0 k
(Table)Tj
2.778 0 Td
(2)Tj
0 g
(\),)Tj
1.2642 0 Td
(indicating)Tj
4.2633 0 Td
(that)Tj
1.7801 0 Td
(the)Tj
1.4684 0 Td
(mean)Tj
2.4491 0 Td
(species)Tj
2.9934 0 Td
(diversity)Tj
3.6453 0 Td
(was)Tj
1.7008 0 Td
(not)Tj
-35.1437 -1.2982 Td
(affected)Tj
3.3278 0 Td
(by)Tj
1.1792 0 Td
(acute-on-chronic)Tj
7.1546 0 Td
(alcohol.)Tj
-10.4654 -1.2983 Td
(Next,)Tj
2.3697 0 Td
(we)Tj
1.3266 0 Td
(examined)Tj
/F14 1 Tf
4.1329 0 Td
()Tj
/F8 1 Tf
(-diversity)Tj
4.4787 0 Td
(to)Tj
1.0262 0 Td
(assess)Tj
2.5228 0 Td
(the)Tj
1.4683 0 Td
(degree)Tj
2.8573 0 Td
(of)Tj
1.0148 0 Td
(dissimilarity)Tj
5.159 0 Td
(of)Tj
1.0148 0 Td
(bacterial)Tj
3.6284 0 Td
(communities)Tj
-32.1957 -1.3039 Td
(between)Tj
3.5262 0 Td
(samples.)Tj
3.6113 0 Td
(These)Tj
2.5625 0 Td
(analyses)Tj
3.4866 0 Td
(determined)Tj
4.8473 0 Td
(no)Tj
1.2755 0 Td
(significant)Tj
4.388 0 Td
(differences)Tj
4.5695 0 Td
(between)Tj
3.5262 0 Td
(pair-fed)Tj
3.4016 0 Td
(and)Tj
-35.1947 -1.2983 Td
(alcohol-fed)Tj
4.6998 0 Td
(mice.)Tj
2.3811 0 Td
(Additionally,)Tj
5.4538 0 Td
(comparisons)Tj
5.3347 0 Td
(within)Tj
2.812 0 Td
(pair-fed)Tj
3.4015 0 Td
(and)Tj
1.7292 0 Td
(within)Tj
2.8119 0 Td
(alcohol-fed)Tj
4.7055 0 Td
(conditions)Tj
ET
Q
q
1 j
1 J
0 w
576 737.1 m
36 737.1 l
36 737.6 l
576 737.6 l
f*
36 741.2598 107.1496 23.6976 re
W* n
q
107.0929 0 0 23.6409 36 741.3165 cm
q
/I0 Do
Q
Q
Q
q
0 0 612 792 re
W* n
1 j
1 J
0 w
7.9999 0 0 7.9999 383.0173 745.7952 cm
BT
/F0 1 Tf
1 TL
-0.005 Tc
0 0 Td
(Alcohol,)Tj
3.7134 0 Td
(the)Tj
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