The barrier functions of crude cervical mucus plugs against HIV-1 infection in the context of cell-free and cell-to-cell transmission.

OBJECTIVE: The cervical mucus plugs are enriched with proteins of known immunological functions. We aimed to characterize the anti-HIV-1 activity of the cervical mucus plugs against a panel of different HIV-1 strains in the contexts of cell-free and cell-associated virus. DESIGN: A cohort of consenting HIV-1-negative and HIV-1-positive pregnant women in labour was recruited from Mthatha General Hospital in the Eastern Cape province of South Africa, from whom the cervical mucus plugs were collected in 6 M guanidinium chloride with protease inhibitors and transported to our laboratories at -80 °C. METHODS: Samples were centrifuged to remove insoluble material and dialysed before freeze--drying and subjecting them to the cell viability assays. The antiviral activities of the samples were studied using luminometric reporter assays and flow cytometry. Time-of-addition and BlaM-Vpr virus-cell fusion assays were used to pin-point the antiviral mechanisms of the cervical mucus plugs, before proteomic profiling using liquid chromatography-tandem mass spectrometry. RESULTS: The proteinaceous fraction of the cervical mucus plugs exhibited anti-HIV-1 activity with inter-individual variations and some degree of specificity among different HIV-1 strains. Cell-associated HIV-1 was less susceptible to inhibition by the potent samples whenever compared with the cell-free HIV-1. The samples with high antiviral potency exhibited a distinct proteomic profile when compared with the less potent samples. CONCLUSION: The crude cervical mucus plugs exhibit anti-HIV-1 activity, which is defined by a specific proteomic profile.

Mucus and Mucins: do they have a role in the inhibition of the human immunodeficiency virus?

BACKGROUND: Mucins are large O-linked glycosylated proteins which give mucus their gel-forming properties. There are indications that mucus and mucins in saliva, breast milk and in the cervical plug inhibit the human immunodeficiency virus (HIV-1) in an in vitro assay. Crude mucus gels form continuous layers on the epithelial surfaces of the major internal tracts of the body and protect these epithelial surfaces against aggressive luminal factors such as hydrochloric acid and pepsin proteolysis in the stomach lumen, the movement of hard faecal pellets in the colon at high pressure, the effects of shear against the vaginal epithelium during intercourse and the presence of foreign substances in the respiratory airways. Tumour-associated epitopes on mucins make them suitable as immune-targets on malignant epithelial cells, rendering mucins important as diagnostic and prognostic markers for various diseases, even influencing the design of mucin-based vaccines. Sub-Saharan Africa has the highest prevalence of HIV-AIDS in the world. The main points of viral transmission are via the vaginal epithelium during sexual intercourse and mother-to-child transmission during breast-feeding. There have been many studies showing that several body fluids have components that prevent the transmission of HIV-1 from infected to non-infected persons through various forms of contact. Crude saliva and its purified mucins, MUC5B and MUC7, and the purified mucins from breast milk, MUC1 and MUC4 and pregnancy plug cervical mucus (MUC2, MUC5AC, MUC5B and MUC6), inhibit HIV-1 in an in vitro assay. There are conflicting reports of whether crude breast-milk inhibits HIV-1 in an in vitro assay. However studies with a humanised BLT mouse show that breast-milk does inhibit HIV and that breast-feeding is still advisable even amongst HIV-positive women in under-resourced areas, preferably in conjunction with anti-retroviral treatment. CONCLUSION: These findings raise questions of how such a naturally occurring biological substance such as mucus, with remarkable protective properties of epithelial surfaces against aggressive luminal factors in delicate locations, could be used as a tool in the fight against HIV-AIDS, which has reached epidemic proportions in sub-Saharan Africa.

Purified human breast milk MUC1 and MUC4 inhibit human immunodeficiency virus.

BACKGROUND: The HIV-AIDS pandemic is prevalent in sub-Saharan Africa. Breastfeeding is a risk factor, with transmission from mother to child being as high as 40%. OBJECTIVES: To determine the antiviral activity of crude breast milk and its purified mucins MUC1 and MUC4 against HIV-1 in patients who were HIV positive compared to those who were not. METHODS: Twenty-one human milk samples were taken from both groups. Breast milk mucins were purified by density-gradient ultracentrifugation in caesium chloride and analyzed by SDS-PAGE, Western blotting and amino acid content. The inhibition of the virus by crude milk and purified mucin was assayed by an in vitro HIV-1 p24 assay. RESULTS: SDS-PAGE for purified mucin showed several high-molecular-weight bands for the HIV-negative group and prominently stained single bands on the stacking gel with faintly periodic acid Schiff-positive glycoprotein bands observed in some cases in the running gel for the HIV-positive mucins. Western blot analysis identified the mucins in both groups to be MUC1 and MUC4. Both mucins showed more intensity on Western blotting for the HIV-positive group. There was no difference in the content of serine, threonine and proline of purified mucins for both groups. HIV-1 was not inhibited by crude breast milk from normal (13/14 samples) and infected individuals (19/19 samples). Fifteen of 20 and 16/18 samples of purified mucin from the uninfected and HIV-positive groups, respectively, inhibited the virus. CONCLUSIONS: Crude breast milk does not inhibit HIV-1, whilst purified mucins do in an in vitro assay.

The characterisation of mucin in a mature ovarian teratoma occurring in an eight year old patient.

INTRODUCTION: The presence of MUC5AC (M1 antigen) and MUC6 have previously been found in ovarian mucinous cyst. We characterized the mucins in the crude mucus and tissue of a mature ovarian teratoma in an 8 year old girl. MATERIALS AND METHODS: Mucins were purified from crude mucus by density gradient ultra-centrifugation in CsCl and analysed by gel-filtration and SDS-PAGE analysis. Mucin identification and expression was by western blotting and immunohistochemistry. RESULTS: Histology showed a tumour with solid and cystic areas, with the cysts lined by colonic and respiratory mucosae. Equal volumes of 'sol' and 'gel' phases of approximately 10.0 ml of crude mucus were obtained. Gel filtration and SDS-PAGE analyses suggested that the mucin was mainly of the large polymeric type which dissociated upon reduction of disulphide bonds with DTT. The colonic and respiratory epithelia predominantly expressed acidic mucin of the sialated and sulphated types respectively. MUC1 and MUC1c were expressed exclusively in respiratory epithelium, MUC2 and some MUC6 (focal) in the colonic tissue and MUC5AC in both tissues. Western blotting confirmed the presence of MUC2, MUC5AC and MUC5B in the secreted gel. Serine, threonine and proline made up the bulk of the amino acids in the sample. DISCUSSION: Ovarian teratoma produced a highly viscous mucus secretion in which the mucin was largely polymeric and of the MUC2, MUC5AC and MUC5B type. The respiratory component of the teratoma expressed MUC1 and MUC1c and the colonic components of the teratoma expressed MUC2 and some MUC6. MUC5AC was expressed in both components.