Is there a Role for Probiotics in our profession?

Probiotics are defined as living

microorganisms that confer a benefit

upon the host

Direct bacteriocidal activity

Lactic acid production

Bacteriocins

Hydrogen Peroxide production

Reduction of energy substrate availability in the gut

Glucose metabolism

Short chain fatty acid synthesis

Protection of epithelial cell (EC) barrier

Inhibition of EC apotopsis

Attenuation of chemically induced EC damage

Maintainence of EC intercellular tight junctions

Reduction of toxins from pathogenic bacteria to the EC

Production of oligosaccharides to prevent attachment of gram negative bacilli to EC

Increased Goblet cell mucus production

Alteration of the host inflammatory response

Direct reduction in pathogenic bacterial load

Direct contact with sub epithelial immune cells via toll like receptors

Activation of Peyers patches to secrete IgA

Changes to Toll like receptor expression in maternal and fetal tissue to support down

regulation of inflammatory processes

Increase in TGF-beta in breast milk

Cochrane Database

Bacterial Vaginosis

Analysis suggests beneficial outcome of microbiological cure with the oral

metronidazole/probiotic regimen and the probiotic/estriol preparation.

Preterm Birth

Although the use of probiotics appears to treat vaginal infections in pregnancy, there are

currently insufficient data from trials to demonstrate any impact on preterm birth and its

complications.

Gestational Diabetes

At this time, there are insufficient studies to perform a quantitative meta-analysis. Further

results are awaited from four ongoing studies.

Neonatal - NEC

Probiotics reduces the occurrence of NEC and death in premature infants born weighing less

than 1500 grams.

Antibiotic related diarrhoea

Based on high-dose probiotics, the number needed to treat (NNT) to prevent one case of

diarrhea is seven (NNT 7; 95% CI 6 to 10).

Microorganis

ms

Cervical Ripening

PTGS-2

PGDH

Preterm Labor

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Fetal Membrane ruptureMyometrial Contractility

Pro-inflammatory Cytokines

Chemokines

Prostaglandins (PGs)

PTGS-2

PGDH???

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??

Anti-inflammatory

Cytokines

??

Toll-like Receptor (TLR-4)

Bacterial Endotoxin (LPS)

Proposed Pathway for Infection/Inflammation

mediated PTB

Pathways to Spontaneous Preterm Birth

• Infection/inflammationproinflammatory cytokines, PG’s ,MMP’s

• Decidual hemorrhagethrombin, MMP’s

• Maternal and/or fetal HPA activationPlacental CRH, 11bHSD2, estrogens

• Pathologic uterine overdistensionCAP gene expression, PG’s, OT

GoldenbGergG et al,

2000

Bacterial Vaginosis

• Change in the microbial ecosystem of the

vagina

• Replacement of lactobacilli by either:

- Anaerobic bacteria (eg. Atopobium,

Prevotella, Gardnerella and Mobilincus);

- Intracellular Mycoplasma

- Aerobic bacteria such as E .coli and

enterococci

Diagnosis of Bacterial Vaginosis

• Nugent Score ( Gold Standard )

• Amsell Criteria

• Culture-based techniques

• Molecular Profiling

Bacterial Vaginosis as a Risk Factor for Preterm Delivery: a Meta-analysis

Leitch, H. et al.American Journal of Obstetrics and Gynecology 189:

139-147, 2003

• Eighteen Studies

• 20,232 Women

• Increased risk of PTB with BV

–O.R. 2.19 (95% CI, l.54-3.12)

Vaginal Microbiome

• > 250 species of bacteria detected using genomic sequencing

• Lactobacillus iners and L. crispatus are dominant organisms in healthy women

• Greater abundance of L.vaginalis, L.crispatus,

L.jensenii and L.gasseri during pregnancy

- Microbiota more stable / less diverse during pregnancy ( Romero , 2014; Aagard, 2012 )

Jean Macklaim, Greg Gloor

Microbial Profiling: Illumina Sequencing

Vaginal Microbiome and PTB

• Increased microbial diversity in women with PTB ( Hyman et al, 2014 )

• Important ethnic influences with a greater diversity in Black women

• No large prospective studies in women at risk for PTB

Probiotics and Human Health

• Preventreatment of Diarrheal Disease

. Treatment of acute infectious diarrhea in

children

. Prevention of antibiotic associated diarrhea

. Prevention of nosocomial and community- acquired diarrhea in children

. Treatment of lactose malabsorption

- Prevention of atopic disease

• Produce H2 O2, lactic acid and/or bacteriocidins which inhibit BV causing bacteria

• Disrupt Gardnerella biofilm surface area, density and depth

• Inhibit adherence of Gardnerella Vaginalis to the vaginal epithelium

Probiotic Lactobacilli in

Human Reproduction

Nutritional Probiotics and PTB

• Norwegian Mother and Child Cohort ( Myhre et al, 2010 )

• Prospective Case Control Study - 950 cases of PTB and 17,938 controls

• Food-frequency questionnaires – assessed intake of milk-based products containing probiotic lactobacilli

• OR for PTB 0.86 ( 95% CI: 0.74,0.99) for those women with high intake of probiotics

Probiotics and PTB• RCT of Lacto for 12 weeks (Krauss-Silva ,2011)

- insufficient sample to make conclusions

• Dietary supplementation with Lacto/Bifidobacterium/Strep – 27 women , Rx or placebo from 33-37 weeks – modulated vaginal microbiota and cytokines (Vitali,2012)

• RCT of Lacto/Bifidobacterium for 14 days before elective C-section- modulated TLR genes in placenta/ fetal gut ( Rautava, 2012 )

Probiotic Lactobacilli and Preterm Birth - Mechanisms?

• Lactobacilli reuteri RC-14 and rhamnosus GR-1 effective in restoring normal vaginal flora in non-pregnant women

• Lactobacilli rhamnosus inhibits pro-inflammatory cytokine production from mouse macrophages

• Central role of cytokines in mediating some preterm labour

Placental Trophoblast cultures

• Placentas collected from healthy pregnant women at term at C-section

• Placental and chorion trophoblast cells separated and grown in tissue culture

• Cells stimulated with LPS

• Pretreatment with Lactobacillus rhamnosus GR-1 Supernatant

Microorganis

ms

Cervical Ripening

PTGS-2

PGDH

Preterm Labor

???

? ? ?

?

?

??

?? (-)

Fetal Membrane ruptureMyometrial Contractility

Pro-inflammatory Cytokines

Chemokines

Prostaglandins (PGs)

PTGS-2

PGDH???

? ? ?

?

?

??

Anti-inflammatory

Cytokines

??

Toll-like Receptor (TLR-4)

Bacterial Endotoxin (LPS)

Proposed Pathway for Infection mediated PTB

Cervical Ripening

PTGS-2

PGDH

Preterm Labor

???

? ? ?

?

?

??

?? (-)

Fetal Membrane ruptureMyometrial Contractility

Pro-inflammatory Cytokines

Chemokines

Prostaglandins (PGs)

PTGS-2

PGDH???

? ? ?

?

?

??

Anti-inflammatory

Cytokines

??

Toll-like Receptor 4)

Bacterial Endotoxin (LPS)

GR-1 SN

Microorganis

ms

Proposed Pathway for Infection mediated PTB

Cervical Ripening

PTGS-2

PGDH

Preterm Labor

???

? ? ?

?

?

??

?? (-)

Fetal Membrane ruptureMyometrial Contractility

Pro-inflammatory Cytokines

Chemokines

Prostaglandins (PGs)

PTGS-2

PGDH???

? ? ?

?

?

??

Anti-inflammatory

Cytokines

??

Toll-like Receptor 4)

Bacterial Endotoxin (LPS)

GR-1 SNGR-1 SN

Microorganis

ms

Proposed Pathway for Infection mediated PTB

Cervical Ripening

PTGS-2

PGDH

Preterm Labor

???

? ? ?

?

?

??

?? (-)

Fetal Membrane ruptureMyometrial Contractility

Pro-inflammatory Cytokines

Chemokines

Prostaglandins (PGs)

PTGS-2

PGDH???

? ? ?

?

?

??

Anti-inflammatory

Cytokines

??

Toll-like Receptor 4)

Bacterial Endotoxin (LPS)

GR-1 SNGR-1 SN

Microorganis

ms

Proposed Pathway for Infection mediated PTB

GR-1 SN reduced LPS-induced PTB

Cytokines Maternal Plasma Amniotic Fluid

LPS GR-1 SN LPS GR-1 SN

IL-1β -

TNF-α

IL-6

IL-12p40 -

IL-10 -

MIP-1α

MIP-1β

RANTES

Cytokines Maternal Plasma Amniotic Fluid

LPS GR-1 SN LPS GR-1 SN

IL-1β - -

TNF-α

IL-6

IL-12p40 - -

IL-10 - -

MIP-1α

MIP-1β

RANTES

Cytokines Maternal Plasma Amniotic Fluid

LPS GR-1 SN LPS GR-1 SN

IL-1β - -

TNF-α

IL-6

IL-12p40 - -

IL-10 - -

MIP-1α

MIP-1β

RANTES

Maternal Plasma Progesterone

n=3-5

Conclusions

Probiotic Lactobacilli rhamnosus GR-1 supernatant is able to:

1) attenuate the LPS – induced increase in pro-inflammatory cytokines and chemokines in placental trophoblast, amnion and decidual cells in vitro

2) Prevent the onset of LPS-induced labour in mice in association with decreased pro-inflammatory cytokines in the absence of changes in maternal progesterone

Conclusions (2)

• Live Lactobacillus rhamnosus GR-1 is able to alter the vaginal microbial profile in the absence of changes in cecal flora

These effects are consistent with L.rhamnosus GR-1 exerting its effect through the secretion of signaling molecules delivered to the intrauterine tissues either systemically or ascending through the cervix.