Pap test, what is it for?

Screening of tumor

It is possible to prevent cervical cancer, and not only, through cervical cancer screening programs implemented on a target female population, aged between 25 and 64, by means of an ethological examination: PAP TEST.
It is called PAP TEST in honor of Papanicolau who in 1941 published the first work where he highlighted the diagnostic value of a vaginal smear to diagnose the carcinoma of the uterus.

Who was Papanicolau?

He was a researcher of Greek origin who had to travel to the USA to continue his studies. Papanicolau had an intuition: by studying the cells of the uterine tumors, with the term "discariosis", he understood that it was possible to distinguish between the cancer cells and the precursors of the cancer cells of the uterus. The dysplasic cells have a bizarre core but are not altered in their entirety. While the tumoral cells not only have a larger and irregular nucleus, hyperchromatic, but they are totally altered with a bizarre shape.

After 1945 a great impetus was given to the value of vaginal smear from the American Cancer Society. The company chose as its medical director his great friend Charles Cameron. One of the first to recognize the potential of the Pap test in reducing cervical cancer mortality. The two met regularly to discuss the work. in short his work became a cornerstone in the prevention of uterine cancer.
These studies were mortaled through studies carried out on the guinea pig and on the wife who daily underwent vaginal cytological samples Papanicolau has also put in place a coloring (Papanicolau coloring) that allows to properly color these smears and see the cellular alterations.

Test procedure

Meanwhile, the abstention from sexual relations is recommended, less than 24 hours from the withdrawal, with or without the condom, because they cause inflammatory changes (from mechanical action), alteration of the bacterial flora, possible presence of exogenous amorphous material. The presence of spermatozoa does not interfere with the evaluation of epithelial cells. In case of cervix with an excess of mucus, blood, inflammatory exudate, the cervix must be cleaned before taking it using a bottle holder to remove the excess material and then a dry gauze pad.

•Do not use wadding: leave residues !;

• Do not wash the cervix with a jet of water or saline: the application of saline solution alters the electrochemical balance of the cells causing artifactual aspects of swelling or breaking of the cellular structures that prevent correct microscopic evaluation. of the Pap test a small amount of cervical cells is taken with a plastic Ayre spatula (not wood) or a cytobrush and a cervical swab. Sampling of the endocervix is performed with a rotation of the Cytobrush of 90-180 °. Further rotation of the brush does not improve the representativeness of the sample and may cause bleeding.

• Only the introduction of the Cytobrush, as it is covered with bristles along its entire circumference, provides a representative sampling of the entire endocervical canal + the rotation of the Cytobrush provides a "deeper" sampling but must not exceed 180 ° because it is too energetic and due to bleeding. The woman is positioned in a gynecological position, the speculum is placed and the spatula is used to obtain a sample of cells. The spatula has a shape complementary to the anatomy of the cervix and once inserted it is able to take cells from the exocervice thanks to a 360 ° rotation; the tampon, on the other hand, quite similar to those used for the pharynx, picks the cells exactly from the endocervix, penetrating the outer uterine orifice. In the conventional pap test, the cells are then streaked on a slide for the laboratory exam. In the liquid phase pap test, a machine prepares a "thin layer" preparation. In the latest generation or "thin" tests, the material is introduced into liquid solutions, to avoid glazing fixation artefacts.
Withdrawals must be made in the various areas, even beyond the transformation region (inside) to be sure about the presence of the cells of the squamocellular junction. Usually the samples are taken in the posterior vaginal fornix and are then "crawled" on a slide that will be analyzed.

Papanicolau only gave a result that was: malignant smear, not malignant, suspected for neoplasm. Only in 1988 following the conference was a new classification proposed which is currently in use, albeit with minor modifications: it is Bethesda's classification.

The Bethesda system has the advantage of having introduced innovations that have allowed to provide indications (there are / there are no malignant cells) in a real diagnostic finding that tells us what kind of injury there is in that particular smear in that particular moment. It has also introduced new terminologies, such as squamous intraepithelial lesion (SIL) which is nothing but the cytological equivalent of CIN.

The term introduced is therefore a synonym for dysplasia and has the advantage of dividing lesions into two groups: high and low-grade (L-SIL and H-SIL, low-SIL and high-SIL), because the ability to distinguish lesions in two groups from a greater reliability than a classification in several groups: if you have to classify anything less "containers" there are and fewer mistakes there are! It also has a clinical value because low-grade lesions can be treated one way, high-grade lesions in another: between the L-SILs, the condylomatous lesions and the CINI are included, among the H-SIL are the CIN2 and the CIN3 and I have told you that most CIN1 regresses, so for the L-SIL a "non-invasive" surveillance will be sufficient while for the H-SIL (cin2 3) it is necessary to deepen the type of surveillance with colposcopy, biopsy samples etc. ..

Another introduction was that of ASCUS, a container where doubtful cases are put in: we keep in mind that we are always analyzing cells that have been broken and not a histological preparation; the detached cells may have alterations that may be dysplastic or other alterations (inflammation etc.) so if there is no certainty these cells are classified in this standby condition that requires further evaluation in different ways (which we will see later).
Obviously the levy must take into account the cycle: it must be done around the tenth-twelfth day of the cycle, then away from menstruation because the blood could cause covers, masks.
The report may be negative for intraepithelial lesions or malignancy but positive for infection; this infection must be indicated and is at the base of the cervical inflammation of the cervix.

Cervicitis is due to the presence in the epithelium of the cervix of a lymphocytic infiltrate, monocytic depending on the pathogen. Among the main microorganisms we remember: Trichomonas Vaginalis, Candida (especially in subjects with immune deficiencies), HSV. We can also find non-neoplastic reports such as reactive cellular changes associated with radiation, intrauterine devices or atrophy.

Types of pathogens repertated during pap smear

Mushrooms (for example, Candida)
Bacteria (for example, Gardnerella)
Virus (for example, Papilloma Virus or HPV)
Trichomonas, a protozoan that causes inflammation
other types of intra-epithelial lesions.

Cells altered at the pap test

A coilocyte is a squamous epithelial cell that has undergone a series of structural changes, which occur as a result of an infection of the cell by the human papilloma virus.

The coilocytes may have the following nuclear alterations:
- Nuclear enlargement
- hypercromasia
- granules of chromatin-citrolols
- variations in the size and shape of the core
Nuclear alterations are accompanied by cytoplasmic "halos", formed, at an ultrastructural level, by perinuclear vacuoles, a cytopathic change caused by E5, a protein encoded by HPV, which is localized in the membranes of the endoplasmic reticulum. Nuclear alterations with the associated perinuclear halo are called "coilocytic atypia".

So there are cells that exhibit alterations that could be interpreted as high-grade lesions, so skip that phase of CIN1; then we have L-SIL, H-SIL and squamous carcinoma of the cervix. Among the glandular we have atypical cells, neoplastic cells and carcinoma. This type of screening based on the recall of women, between 25 and 64 years, every 2-3 years has led today to encouraging results: in the last 30 years in Italy the carcinoma of the cervix has been reduced by 2/3. The PAP test, or the evaluation of the smear, represents the most powerful screening system currently existing in oncology, this is because it is based on two facts: women are called periodically in the critical period of onset and accuracy of the test. From 20 / 100,000 cases to 7 / 100,000 cases in the last thirty years.
Let's go back to the Coilocyte we saw earlier: it represents the alteration determined by the HPV virus, they are atypical cells that have a perinuclear halo.

HPV infection

Can cervical carcinoma without HPV infection exist? In fact no, it has been seen that HPV and carcinoma of the cervix are closely linked: in all carcinomas the presence of the virus is evident. The virus was discovered with the advent of PCR.
Viral infection is extremely widespread: 80% of the sexually active female population contract the virus at least once, the infection persists for a variable time (about 2 years) and then disappears; this explains why the CIDs disappear because the effects are related to the virus. Only a portion of the infections become persistent due to the deficiencies of the immune system of the PC, of the smoke etc .. and therefore the viral genome integrates into the genome of the host cell and begins to give rise to the transformations at the base of the low-grade CID, the which in most cases regress within 5 years; after that if the viral integration persists the CID becomes of high degree, here too regression up to 70% of the young and up to 30% of the women over fifty (because in the juvenile population the virus is much more frequent), to the end of the path we get to the invasive carcinoma. As I told you to get to the carcinoma it takes tens of years.
HPV has a tropism for the epithelial cells of the mucous membranes so we can also find it at the level of other districts, such as the oropharynx, but here the infection / neoplasm association is much less strict, other factors will be indispensable. There are several species of HPV:
. a group that affects the mucous membranes of the genital tract
. a group that affects the skin, responsible for the appearance of warts
. a group that interests animals
The first group is certainly the most important; in each group we find different viral types grouped in species.
The viral types found are more than 100 (30 those of the genital tract) and we can divide them into genotypes with low oncogenic risk (associated with benign lesions) and genotypes with high oncogenic risk. Here, however, there is confusion because it is thought that the viruses at high risk of cancer is the one that gives cancer, but NO: the infection that I said there is the vast majority of people affects all types, infections can be single or multiple.
On the 6th and 11th, those types are linked more to condylomatous lesions, on the 16th and the 18th they are linked to high-grade lesions.
The virus is a circular DNA (7900 base pairs), has a capsid produced by the L genes, has tropism for epithelial cells. The capsid consists of two structural proteins LI and L2 and other non-structural proteins E (from El to E7), responsible, among other things, for the cytopathic effect of the virus (especially E6 and E7), interference in the binding with cytokeratins (E4). The virus does not cause cellular necrosis but once internalized in the cell it replicates, is assembled and released with the Early (E) and Late (L) proteins. The oncogenic power of the virus is bound to the E6 and E7 proteins as they interfere with two tumor suppressor genes: p53 and RB (the retinoblastoma gene).
When the virus integrates it causes a breakdown at the level of E2, which has the function of regulating the virus in the expression of E6 and E7, then breaking in E2 the virus no longer has control of the regulation and produces E6 and E7 in abnormal way. This, in the infected cell, determines an interference in the normal function of "e2f", transcription factor regulated by the RB gene, and at the same time of p53 which, thus altered, causes a non-blocking in G1 of the cell, which therefore continues to proliferate. with an altered genome.
Obviously the altered genome can not be repaired, errors accumulate and this "genomic instability" will then lead to cancer, all this during DECENNI, it is not an immediate thing. It is important to distinguish transient infections from persistent infections, and also the type of virus contracted. The various steps that we have seen are conditioned by the body's defenses (a defiled piece has a different situation compared to a healthy person), age, sexual relations, smoking, the immune system, use of oral contraceptives (which have an action favoring proliferation). More than 70% of cancers have HPV infection 16 and 18 (which are ubiquitous), although other viruses have been found. Immagine correlata

Until a few years ago, and even now in some parts, the HPV TEST is proposed in the triage of ASCUS lesions. We said that if we have an ASCUS we do not know what it is -> let's do the HPV TEST -> If the test is positive it means that these lesions are dysplastic and we do the colposcopy; if the test is negative, it must be repeated within 6 months.
Currently in Italy the HPV TEST is foreseen in three conditions: triage of the borderline ASCUS cytology in the primary screening of experimental studies in the case of treated pts (to see if the virus persists or not)

In the triage of borderline cytology the PAP test has a variable sensitivity, not high, while the HPV TEST has a very high sensitivity: then this test is important not because there is an infection but if it DOES NOT 'the infection or who does not has the infection can feel comfortable (has negative predictive value); who has the infection does not mean anything!
It has recently been proposed to overturn the situation: first use the HPV TEST, which is more sensitive and less specific, to select all infected people and then this group is subjected to triage with cytology, which is more specific. This is a guideline now used in America, in Italy instead we do it experimentally only in two regions but has not yet been extended to the whole nation.

HPV is contracted by women in 80% of the female population during their lifetime; 46% have a high probability of contracting the virus three years after their first sexual relationship, even though they only had one relationship. So the concept "+ sexual intercourse = + possibility of contracting the virus" is true but up to a certain point! In fact, the virus can, albeit less, be contracted even with contaminated objects that come into contact.
There is a quadratic anti-HPV vaccine (anti 6-11-16-18) also proposed free from the Sicilian region to adolescents to prevent significant damage; remember, however, that the vaccine only covers 1/3 of the viruses circulating.

kwokn more >> lesioni tumorali e pap test

tumor index