α-Defensins were detectable among most (79%) of the 1-week breast milk samples tested, and HIV-positive women with higher concentrations in their breast milk were less likely to transmit HIV to their infants via the intrapartum or postnatal route than women with lower concentrations, even after adjusting for the quantity of cell-free virus in the milk and for maternal CD4 cell count. A recent study among uninfected women in the United States also observed that α-defensins are detectable in breast milk from most women.8
Thus, α-defensins seem to be yet another anti-infective factor, among the plethora of immunologically active components,11
produced in human milk under certain circumstances. Concentrations were higher in the US study8
than in ours, possibly because of population differences, differences in postnatal ages at which the samples were collected, or differences in laboratory methodologies. Nevertheless, within our study population, higher concentrations of α-defensins distinguished the nontransmitting from the transmitting HIV-positive women.
α-Defensins tended to be detected more often and at higher levels in the breast milk from HIV-positive mothers compared with milk from HIV-negative control mothers. Furthermore, among HIV-positive mothers, α-defensin concentrations were correlated with cell-free breast milk viral load (ie, as HIV RNA quantity in milk increased, so did α-defensin concentrations). Thus elevated α-defensin concentrations may be, at least in part, stimulated by active HIV infection. α-defensins may be produced to augment mucosal immune defenses in the breast when physiologically needed.
The amount of HIV RNA quantified in breast milk was a strong predictor of mother-to-child HIV transmission in our data, as has been previously observed in other studies.12,13
Paradoxically, despite the correlation between breast milk viral load and α-defensins, once breast milk viral load was controlled for, α-defensin concentrations were strongly and significantly associated with lack of HIV transmission. In other words, at comparable breast milk viral load levels, elevated α-defensin concentrations were associated with a reduced risk of HIV transmission among breastfed infants. A limitation of our study is that we did not measure cellassociated virus in breast milk, which has been suggested to be a more important predictor of postnatal transmission than cellfree virus.14
Failure to adjust for this parameter is unlikely to explain our observed association between α-defensin concentrations and lack of transmission, however, because the quantity of cell-associated virus is significantly correlated with the quantity of cell-free virus14
; adjustment for cell-free virus in our data strengthened rather than diminished the magnitude of the association. It is interesting to note that in the study of cell-associated virus, levels in breast milk samples collected soon after birth were not associated with postnatal transmission14
and that, in general, innate immune factors, including α-defensins, tend to decline from high levels in colostrum to lower levels in samples collected at later postnatal ages.8
The significance of this is unclear and needs to be further investigated.
Lower CD4 cell counts were independently associated with transmission in our data possibly because of more robust maternal adaptive immunity. Declining CD4 cell counts tend to correlate with increasing deficits in antigen-specific cell-mediated immunity.15
In our data, however, immunosuppression (measured by a low CD4 cell count) was not associated with lower levels of α-defensin concentrations in breast milk and thus did not explain the association we observed with transmission.
These results are consistent with the studies among HIV-infected adults that have observed that high concentrations of α-defensins are correlated with long-term nonprogression and are observed among HIV-exposed high-risk but uninfected individuals.4,7
Our data suggest that α-defensins may also have some role in prevention of HIV transmission to breastfed infants. Because all infants in our study were breastfed, a limitation is that we cannot distinguish whether the association pertains only to postnatal transmission through breastfeeding or also to intrapartum transmission during delivery. Both of these routes of transmission involve exposure to maternal mucosal fluids,16
and it would be interesting to investigate whether α-defensin concentrations are similarly elevated in cervicovaginal secretions among nontransmitting HIV-positive mothers.
It is not a novel suggestion that maternal breast milk factors may influence infant susceptibility to infection. Breast milk is known to contain several immunologically active components that augment the newborn’s immune defenses.11
Several of these factors, including secretory leukocyte inhibitor (SLPI), lactoferrin, lysozyme, human b-defensin, and regulated upon activation, normal T-cell expressed and secreted (RANTES) among others, also have anti-HIV activity under certain conditions. A limitation of our study is that we cannot ascertain whether the elevated α-defensin concentrations are simply correlates of other factors in breast milk with anti-HIV activity that might be present under the same conditions that promote local α-defensin production. Studies have not necessarily observed significant associations between concentrations of these factors in breast milk and reduced HIV transmission17,18
; thus the relative importance of these factors in reducing the risk of postnatal transmission remains unknown.
Elevated α-defensins or other innate immune factors in breast milk may influence transmission by processes involved in reducing the infectivity of virus in breast milk and/or in increasing infant resistance, possibly via mechanisms analogous to passive immunization. They may also support adaptive immune responses operating locally in the breast or in the child. Our study is unable to determine whether the association between elevated α-defensin concentration and reduced transmission is simply a result of the correlation between this innate immune factor and other immune responses in breast milk, such as neutralizing antibody responses, HIV-specific cytotoxic or helper cell responses, or natural killer cells. Some but not all studies have observed associations between anti-HIV antibodies in breast milk and reduced transmission.19,20
HIV-specific cytotoxic cells also seem to be quite common in milk from HIV-positive women.21
α-defensins may be one of perhaps several mucosal immune factors or responses that are stimulated in the breast and may help to explain why, despite the prolonged and extensive exposure of breastfed infants to virus, transmission through this route is relatively inefficient.
These data raise the question of why some mothers produce high levels of breast milk α-defensins when their breast milk viral load is high, whereas others apparently do not. Genetic polymorphins that facilitate production of these antiviral factors may be involved and have been identified for b-defensins but not for α-defensins.22
Our data suggest that there may be a role for α-defensins in prevention of mother-to-child HIV transmission. Further investigation of the dynamics and source of this mucosal antiviral factor and its association with other local and systemic immune factors and with HIV transmission through different routes is warranted.