Epithelia, tissue and growth disorders

Epithelia, tissue and growth disorders

Certain cells of the body are specialised for specific functions. A tissue is defined as a group of cells with a similar structure and function, there are 4 main types of tissue:

1. Epithelium
2. Connective tissue
3. Nervous tissue
4. Muscle

Epithelia

Epithelia are derived from 3 layers, referred to as the germ layers; the ectoderm, mesoderm and endoderm. They line and cover all of the body's surfaces with the exception of articular cartilage, tooth enamel and the anterior iris surface. Its basic functions, dependent on its location are: 

• Protection: for the skin
• Absorption: for the small and large intestines
• Transport: on the surface mediated by cilia cells
• Secretion: for glands
• Excretion: for the tubules of the kidney
• Gas exchange: for lung alveolus
• Gliding: between surfaces, for the mesothelium

Once fertilisation occurs, the sperm and egg form a zygote, then grow into a blastocyst, and then become what is referred to as a gastrula. The ectoderm, the external layer of the gastrula, derives epithelia such as skin cells, brain neurones and cell pigments. The mesoderm, the middle layer of the gastrula, derives cardiac muscle, skeletal muscle, kidney cell tubules, red blood cells and smooth muscle. The endoderm, the internal layer of the gastrula, derives the lung alveolar cells, thyroid cells and pancreatic cells. Germ cells are also derived from the gastrula, these make up sperm cells in males and egg cells in females. Sometimes however these cells can end up in random places, causing disfigurements, such as teratomas. Germ cells are totipotent, meaning they can form anything derived from the germ layers. 

Due to their location and functions, epithelial cells are renewed continuously by mitosis, this however makes them prone to causing cancers. They lack a direct blood or lymphatic supply, thus their nutrients have to be provided via diffusion. There is almost no free intercellular substances. The structural organisation of the epithelium is maintained by cell adhesion molecules and junctional complexes. The cells are anchored to a basal lamina, this and connective tissue components interact in oder to form the basement membrane.  The cells have a structural and functional polarity, apical and basal (top and bottom), however in cancer, this polarity is lost. 

Epithelia classification

This classification is based upon the cellular morphology, not the cellular functional characteristics. It is determined by their shape and arrangement. 

Shape can be:

• Squamous: very flat, almost 2 dimensional squares
• Cuboidal: 3 dimensional squares, cubes
• Columnar: 3 dimensional columns, rectangular

Arrangement can be:

• Simple: single layer
• Stratified: multilayered
• Pseudo-stratified: technically a single layer, but looks double as cells are in a top and tail position
• Transitional: multilayered however can be relaxed and distended

Figure 1

Simple epithelium 

Simple squamous is a single layer of flat cells, and usually forms membranes. It lines body cavities and lungs and capillaries. It is given the name endothelium when it lines the blood and lymphatic vessels, and mesothelium when lining the body cavities. Due to their thin nature, they're useful for gaseous exchange in the alveolar. 

Simple cuboidal is a layer of cube like cells, they're common in glands and ducts. They also form the walls of kidney tubules, and cover the ovaries. This is a large contributor to the incidences of ovarian cancer, a theory suggests that every time an egg is released, it may rupture the outside of the ovary where it will then have to heal, thus the longer the ovulation period, an increase in the chance.

Simple columnar is a single layer of tall, rectangular cells. They often include goblet cells which secrete mucus, they also line the digestive tract. 

Pseudostratified is a single layer of cells, however some are shorter than others giving it the appearance of a double layer, sometimes it can be ciliated, this occurs in the respiratory tract. They may also have a absorption or secretory function. 

Stratified epithelium

Stratified cuboidal cells are multilayered, and they ave an apical surface (the top of the top layer). It is rare if surface cels are cuboidal, this only occurs in sweat gland ducts and the male urethra.

Stratified squamous cells of the free edge are flattened, they're usually found as a protective covering where friction on the area would be common, such as the skin,mouth and oesophagus. These can be keratinised also. 

Transitional epithelium

Shape of the cells will depend upon the amount of stretching required, these kind of cells line the urinary system.

Microvilli

These are finger-like processes which extend from the apical surface of cells, usually around 1-2micrometres in length. Their main component is their core of actin filaments, which are around 6nm thin, these are named microfilaments. They're very extensive in any absorptive epithelium, as they increase the cells surface area, and can be referred to as a striated border or brush border. Bundles of actin filaments extend into a part of the cell which is named the terminal web (see figure 2), these are actually anchored to the cell.

Figure 2.

Cilia

Cilia are long cytoplasmic extensions around 5-10micrometres in length. These do not contain actin filaments. Instead they have a complex  arrangement of microtubules (9 around the edges and 2 in the centre) named "axoneme", which have small appendages named dynein arms which allow for movement, they're essentially molecular motors, thus the main component of cilia is tubulin. Cilia facilitate flow of fluid over an epithelium, for example on the trachea, a tubular organ. The structure is the same as flagellum (see fig 3). 

Figure 3.

Kartagener's Syndrome (KS)

This autosomal recessive disease is essentially immotile-cilia syndrome. It occurs in around 1 out of 32,000 live births in the US. The symptoms include chronic upper and lower respiratory tract disease, as a result of ineffective mucociliary clearance, sterility in males, as the flagella in the sperm doesn't work properly and approximately 50% of those suffering with the syndrome display situs inversus, which is where the visceral internal organs are mirrored (different sides to normal), also referred to as transposed viscera. The biopsy examinations reveal abnormal/non motile cilia. The axoneme is lacking in dynein arms.

Basement membrane

This is an extracellular component which is in direct contact with the basal domain of epithelial cells, it can be made visible under a light microscope with the use of a periodic acid-Schiff (PAS) stain, making it PAS positive. The basement membrane can be defined into 2 layers/laminae:

1. Basal lamina: this contains laminin, fibronectin, type IV collagen, heparan sulfate proteoglycans and nidogen.
2. Reticular lamina: contains type 3 collagen.

The main components of these 2 layers are glycoproteins. They can be witnessed under the electron microscope.

Basal lamina functions

The basal lamina as a variety of functions:

• Adhesion of epithelial cells to underlying CT, in skeletal muscle the BL maintains the integrity of the tissue, and any disruption to it will cause muscular dystrophies.
• Act as a selective permeability barrier, with the filter being based upon the charge of the molecule and its size.
• Good substrate of cellular migration, for example during wound repair or embryonic development.
• As a barrier critical to metastatic potential in epithelial cancer cells referred to as carcinomas. A carcinoma which is in situ means it has not breached the basement membrane. 

The main protein making up the BL is type 4 collagen, this forms flat sheets and has the ability to self polymerise. Perlecan also has the ability to self polymerise, and laminin also has the ability to self polymerise and also can interact with itself.

Secreted protein acidic and rich in cysteine (SPARC)

This is also known as osteonectin.  It is expressing during many stages of development in a variety of organisms, however it is restricted in adult vertebrates primarily to tissue which under consistent turnover or remodelling, such as the bone. The over expression of SPARC is found in various human malignancies. 

Melanoma cells with suppressed expression of SPARC could no longer generate tumours, however melanoma cells which induced expression of SPARC has 100% tumorigenicity. High SPARC levels are also associated with metastatic tumours, as it induces the expression of MMPs, MMP1, MMP2 and MMP9, which are involved in the degradation of basement membranes and extra cellular matrix. This is crucial process for invasion and metastasis in human malignancies. 

Metastatic cascade

Tumour cells coat themselves in the host platelets in order to avoid detection from the immune system. Collagen is an example of an angiogenesis inhibitor. Arrestin, canstatin, turnstatin and endostatin, all type 4 collagen have specific anti-angigenic properties. Without angiogenesis, a metastatic deposit will only grow around 2mm before dying. If the integrin binding it is inhibited, then so it growth of the tumour. 

Cadherins 

The name is derive from the term "calcium-dependent adherence protein", the family consists of around 90 members, whom al participate in the interactions between cells of the same type, which is ideal for epithelial layers. These interactions which connect the plasma membranes of adjacent cells, forming 2 cellular junction types:

1. Zonula adherens: these are small, spot like junctions located near the apical surface of epithelial cells 
2. Desomosomes: stronger and more extensive junctions, present in epithelial and muscle cells. 

Linkage of cadherins with the cytoskeleton occurs through 2 classes of catenins; beta-catenin links cadherins with alpha-catenin which in tern connects to actin, thus completing the connection with the cytoskeleton. 

E cadherins

These mediate homotypic adhesions in epithelial tissue, thus serving to keep the epithelial cells together, and to relay signals between the cells. In several epithelial tumours, e.g adenocarcinomas (gland cancer) of colon and breast, E-cadherin is down regulated, this reduces the cells ability to adhere to one another and promotes their detachment from the primary tumour, and also their advance into other tissues. In some tumours, E-cadherin is normal, but its expression is reduced due to mutations for one of the catenin genes. 

Epithelial-mesenchymal transition (EMT)

This is the name given to the process in which epithelial cells lose their polarity and cell to cell adhesion, and gain invasive and migratory properties, turning them into mesenchymal stem cells, a multi-potent type of cell which has the ability to differentiate into many cell types. 

EMT is crucial in a number of developmental processes, including mesoderm formation, neural tube formation, and foetal development (embyrogenesis). It is also found to occur in wound healing, and in the initiation of metastasis for cancer progression. Normal cells have contact inhibition, or spatial awareness, however cancer cells do not. The cells have an amoeba shape, they detach and become invasive and motile, they break through the basement membrane and reach the blood vessels in order to move to another location. Once they have found a place to form and grow, the process is reversed by a process called mesenchymal-epithelial transition. In order to check for this, a stain can be carried out which will show that any mesenchymal cells will not contain E-cadherin. 

When the above markers are switched on, the epithelial marks seen above in green are turned off, and the mesenchymal markers seen above in red, are turned on. Zeb 1 and 2 should also be in the list, as they are recently discovered to be down regulators of E-cadherin. These can technically be considered as growth factors as they trigger a signalling mechanism. 

Cellular adaptation

Adaptive responses result in an increased tissue mass, the increased functional demand or endocrine stimulation are what usually cause hypertrophy (increase in volume) and hyperplasia (increase in cell number). These new patterns of growth are stable whilst the causative stimulus persists, however once it is removed, the tissue will return to normal growth patterns. 

Size

Acquired changes may be physiological or pathological, this is due to external influences, however these can potentially be reversible.  

Atrophy: this is a reduction in size or activity. Examples of a physiological atrophy are the thymus as it ages, a thymus in a child is much more larger and dense in comparison to a thymus in an adult, ovaries after menopause and immobilisation of a broken limb. An example of a pathological atrophy are denervation, loss of trophic hormone, reduced blood supply and excessive pressure. 

Hypertrophy: this is an increase in cell size. Examples of a physiological hypertrophy are increased skeletal muscle bulk due to training, or the uterus during pregnancy. A pathological example of hypertrophy is cardiac muscle due to an abnormal increase in workload, e.g. aortic valve stenosis. Mechanism sensors appear to be the major triggers for physiological hypertrophy, and agonists and growth factors are more involves in the pathological hypertrophies. 

Hyperplasia: this is an increase in the number of cells. Physiological examples, which are usually hormone induced, include adrenal cortex due to stress, breast enlargement due to puberty and pregnancy, may coexist with hypertrophy such as the uterus in pregnancy. Pathological examples which can also be hormone induced, include the adrenal cortex due to a pituitary tumour secreting ACTH, enlargement of the prostate and endometrial hyperplasia, this could be considered as pre neoplastic is glands are atypical. There is also compensatory hyperplasia, for example in liver and kidney resection. 

On the left, is a normal uterus and on the right is an enlarged uterus during pregnancy. 

Epidermodysplasia verruciformis

This is an example of hyperplasia driven by a viral infection, in this case, HPV is the cause. There are obver 150 types of HPV, the most commonly known are 16 and 18 which cause cervical cancer, and 6 and 11 which cause warts. HPV 5 and 8 cause growths (seen in the photo below), others do also but these are the main contributors, the way they have affected 

this man does suggest he was in an immunocompromised state. It is currently being researched how zinc and EVR1 and EVR2, genes which code for a zinc transport mechanism, affect HPV.  

Metaplasia

This is a reversible change in which one cell type, epithelial or mesenchymal, is replaced by another cell type. This represents an adaptive substitution of cell types which are sensitive to stress with cells that are better to be able to withstand the different environment, which would cause stress or trauma on the original cell. This does not result from a change in the phenotype of a differentiated cell type, instead it is the result of a reprogramming of stem cells which are known to exist in normal tissues, or of undifferentiated mesenchymal cells present in connective tissue. In metaplastic change, these precursor cells differentiate along a new pathway. For example, the metaplastic transformation of oesophageal stratified squamous epithelium into mature columnar epithelium, which can be found in intestinal cells that cause acid reflux this is a pre neoplastic condition, this metaplastic change can develop into cancer if the stress/trauma to the cells is not removed. A risk factor towards this is smoking, although it is primarily caused by HPV, HPV itself can cause a variety of cancers, including cancer of the vulva, penile, oral, anal and skin cancer. HPV requires the presence of stratified squamous cells in order to grow. 

Dysplasia

This is the term used in order to describe cells which have:

• An increased rate of cellular division often with abnormal mitotic figures
• Incomplete maturation
• Loss of normal cellular architecture
• Tend to have a high nuclear to cytoplasmic ratio

The presence and severity of dysplasia can be regarded as an indicator of the risk status of a precancerous lesion, and can be graded however this is very subjective, and so is basically the opinion of the the scientist observing it. Severe dysplasia indicates a very high risk of the subsequent development of cancer. Pre-neoplastic cells however can sometimes be reversible. Dysplasia is defined by 19 parameters.

Scrapes can be taken of the cells, however these are usually only on the surface of the epithelial layer. In cervical cancer, they can be categorised into cervical intraepithelial neoplasia 1, 2 and 3. 

Histological changes in dysplasia diagnosis

• Loss of polarity of basal cells
• The presence of more than one layer having a basaloid appearance
• Drop-shaped rete-ridges (epidermal thickenings)
• Increased nuclear-cytoplasmic ratio
• Nuclear hyperchromatism
• Enlarged nucleoli
• Increased number of mitotic figures
• Mitotic figures that are abnormal in form
• The presence of mitotic figures in the superficial half of the epithelium
• Cellular and nuclear pleomorphism (variability in size and shape)
• Irregular epithelial stratification
• Loss of intercellular adherence 
• Keratinisation of single cells or cellular groups in the prickle cellular layer
• Mild and severe dysplasia is easier diagnose than moderate
• If any of these changes become extreme, it can lead to cancer
• In cancer you get more than one nucleoli

There are 3 major problems attached to the importance of epithelial dysplasia in predicting malignancy development;

1. There is a set of criteria for assessing epithelial dysplasia however the overall diagnosis is very subjective 
2. Not all lesions showing dysplasia will become malignant, some regress
3. Carcinomas can develop from lesions in which epithelial dysplasia has not been diagnosed in previous biopsies.

In this case, there is a substantial requirement to improve the histological assessment of epithelial dysplasia, or, since it does not seem to be invariably associated with or even a necessity for malignant development, it may be necessary to develop other methods for predicting the malignant potential for premalignant lesions. 

DNA ploidy analysis is a good prognostic marker, as it is objective, in comparison to histological grading, which is subjective.

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Cervical cancer

Cervical cancer

Figure 1. Picture provided by Roy Stewart - Cervical cancer lecture presentation.

The area joining the cervix and the vagina is the main place where cancer can form. 

The vagina has a slightly acidic environment, and this can be very harmful to the cells found on the ectocervix (also referred to as the exocervix), during puberty the cervix opens, thus in order to deal with the acidity, the cells must undergo metaplasia (genetic reprogramming of the basal cells), causing a migration of the squamocolumnar junction into the transformation zone. As a woman ages, the squamocolumnar junction becomes restored to its original site. The squamocolumnar junction and the transition zone is where most things occur, in a smear, it is required to have endocerivcal and ectocervical cervical cells, along with parts of the transformation zone.  

Cervix histology

The cervix can be divided into 2 parts:

1. Ectocervix: visible to the naked eye on a vaginal examination, it is covered by a stratified non keratinising squamous epithelium continuous with the vaginal vault. The squamous epithelium converges centrally at a small opening termed the "external os". In a woman who hasn't completed a pregnancy beyond 20 weeks (deemed as nulliparous), the os is virtually closed. 
2. Endocervix; just anterior to the os, it is lined by simple columnar, mucus secreting epithelium that dips down into the underlying stroma to produce crypts (endocervical glands).

The point of these two areas meeting is referred to as the squamocolumnar junction, its actual position can vary due to the cervical anatomy and the basal cell and subcolumnar reserve cell distribution that occurs. The progressive differentiation of these basal/reserve cells which governs the microanatomy of this region, thus resulting in migration of the SJC. The area of columnar epithelium which is replaced by squamous epithelium is referred to as the "transformation zone", it's within here and the SJC where precancerous lesions and squamous carcinomas develop.  

Human Papillomavirus (HPV)

HPV is a member of the Papoviridae virus family, and there are over 100 types of HPB which all have the ability to affect different epithelial surfaces, these include the hands and feet, as well the genital region. Some HPV types are the cause of warts and also genital warts (condyloma acuminata), HPV types 5 and 8 are associated with the development of "tree man syndrome", whereas some are the causative agent in cervical cancer. HPV has also been associated with other types of cancer, such as oral, laryngeal, anal and skin cancer in patients whom are immunocompromised. HPV types such as type 16 and 18 associated with cancer are referred to as "high-risk" HPV types, whereas those associated with benign warts, such as HPV types 6 and 11 are referred to as "low-risk" HPV types. The HPV vaccine given out to young girls in recent years targets the HPV type 16 and 18. 

Figure 2. Picture provided by Roy Stewart - cervical cancer lecture presentation

The viral capsid (fig 2) consists of 72 capsomeres, each of which is composed of 2 viral proteins, L1 (major capsid protein) and L2 (minor capsid protein). HPV uses a circle of double stranded DNA for a genome, which is around 8000 bp in length. All of the proteins the virus uses are encoded for within one strand of the DNA, thus are all read in the same direction. The genes within the DNA can be categorised into late and early, affiliated with when they're expressed during the course of an infection. 

The virus infects the basal cells of the epithelium through minor abrasions in the skin, these basal cells then divide, and their progeny will also contain HPV DNA. During the early stages of the initial infection, the number of viral genome copies is small, around 50-100, and the genome exists as an extrachromosomal DNA circle, also referred to as an episome, which goes on to replicate in the same manner as the host cell chromosomes.

The viral genome doesn't undergo amplification until the basal cells begin the differentiation process. As the cells approach terminal differentiation, the late genes L1 and and L2 are activated, and transcribe the major and minor viral capsid proteins. At this point, the viral genome number will have increased rapidly, so that each cell will produce 1000s of viral particles. As the cells approach the surface of the skin, they're shedded, and the virus particles are released in order to infect other cells and spread to other hosts. 

When the virus integrates into the hosts DNA, this is when a malignancy can arise. The plasmid linearises by losing E2 and E4, these are involved in controlling the virus and are no longer required once inserted into host DNA. With the loss of these control genes, the others become switched on. The main problem genes associated with malignancies are E6 and E7, and their regulation will become heightened. 

Figure 6.Picture obtained from Roy Stewart - Cervical cancer lecture presentation

Low grade: as cells grow and mature, they make new viruses, however as it is an episome, it doesn’t switch on the E6 and E7 genes, however there is a partial turn over of E6 and E7.

High grade: E6 and E7 switched on with no control, they are produced in a ridiculous number.

Figure 7. Picture obtained from Roy Stewart - Cervical cancer lecture presentation

It is thought that as an average estimate, cervical cancer takes around 10-15 years to develop fully. A main point is that a persistent infection is required in order for the development of neoplastic change to occur. Other cofactors are involved with the progress of the disease such as smoking, oral contraceptions etc. 

Cervical intraepithelial neoplasia (CIN)

A persistent infection can be categorised as CIN 1 (mild dysplasia), this infection can be cleared easily, however if left, it may progress into CIN 2 (moderate dysplasia) or CIN 3 (severe/invasive dysplasia), and alongside other cofactors the viral DNA will integrate itself and this can progress into cervical cancer. A screening will be able to pick up any preneoplastic changes, hence their importance.

Figure 8. Picture obtained from Roy Stewart - Cervical cancer lecture presentation

HPV binding and cellular entry

The viral particle interacts with the cellular surface it is trying to infection, through interaction between the L1, major capsid protein, and heparan sulfate proteoglycans. There is evidence which suggests that a secondary receptor is involved, possibly providing a role for the L2, minor capsid protein in interactions on cellular surface. HPV entry in intro is carried out through the binding to a cell surface receptor, whereas in vivo the basement membrane is identified as the virus binding primary site. This binding (in both scenarios) promotes a conformational change, affecting both L1 and L2 proteins, enabling them to interact with an uptake receptor. Most HPV types enter the cell via an endocytic mechanism which is clathrin-dependent. Clathrin is a protein which is associated with coated vesicle formation.  This process occurs slowly and also not at the same time, it can be associated with a long residence on the cell surface. Many details of this process however are yet to be clarified.

Host DNA integration

When the virus integrates into the hosts DNA, this is when a malignancy can arise. The plasmid linearises by losing E2 and E4, these are involved in controlling the virus and are no longer required once inserted into host DNA. With the loss of these control genes, the others become switched on. The main problem genes associated with malignancies are E6 and E7, and their regulation will become heightened. 

Malignancy

Figure 3. Picture obtained from Roy Stewart - Cervical cancer lecture presentation

Within different stages of the cell cycle, different cyclins are made, and they become activated when in the presence of their specific cyclin dependant kinase (CDK). The retinoblastoma protein (Rb), a tumour suppressor family protein, is also has a crucial role in controlling of the cell cycle. It tightly binds to transcription factors of the E2F family, and represses the transcription of particular cell cycle genes.

In the G1 phase, cyclin D becomes activated by CDK4,6, which has the ability to phosphorylate Rb, causing it to become inactivated. Cyclin E is released and activated by CDK 2, this then phosphorylates the Rb further, causing it to become hyperphosphorylated. 

Figure 4. Picture obtained from Roy Stewart - Cervical cancer lecture presentation

E7 protein  latches onto Rb via the CR2 domain, accelerating the cell into a premature state of S phase by disrupting its binding to the E2F molecule, E7 has a higher affinity for Rb than E2F thus causing its displacement. This oncoprotein is involved in both differentiation and proliferation, suggesting it can uncouple these processes  through transcription of different gene subsets. This causes eventual degradation of Rb, and as a consequence a lack of regulation within the cell cycle, promoting cellular proliferation and growth, these are hallmarks of cancer. 

E7 also affects cyclin E by deregulating its transcriptional levels, this is caused by a decrease in repression. E7 also interferes with the TGF beta tumour suppressor pathway (Smad 2,3,4), inhibiting its function.

Cyclin A alongside CDK2 becomes assembled during the S phase, and will remain active though the entirety of G2 phase also.

Figure 5. Picture obtained from Roy Stewart - Cervical cancer lecture presentation

The p53 protein is a tumour suppressor protein which regulates the cell cycle between the G2 stage and M stage of the cell cycle (the G2/M checkpoint), and is tightly regulated through MDM2. Usually, if there is an error in the DNA of a cell, the p53 protein will inhibit it's proliferation and cause the cell to carry out apoptosis. However in the presence of E6, this does not occur. E6 binds to a ubiquitin ligase called E6AP, when this E6-E6AP complex comes into contact with an p53 protein, it interacts with it and causes it to become ubiquinated. Once this happens, the p53 protein is then sent for destruction via the proteosome pathway. As the protein is degraded, it's function cannot be carried out, thus allowing damaged cells to continue through the cellular cycle and proliferate in an unregulated manner. 

E6 has another role in the up regulation of hTERT, alongside c-myc, which generates telomerase. If a loss of telomeres occurs, and a significant amount of replication occurs, the chromosomes may "stick together", if these chromosomes are then replicated again, the chromosomes may fragment. When a cell detects a shortage in telomeres, it goes into a coma like state, a cancer prevention mechanism, referred to as senescence. Germ cells do not get a shortage of telomeres, this is because their telomere mechanism is always switched on, so as to not have any anomalies in sex cells. The production of telomerase essentially expands the life cycle of a cell, thus the up regulation will cause the cell to become "immortal".  

P16 is another tumour suppressor, it's role in the cell cycle is to inhibit cyclin-dependant kinases (mostly CDK2 and CDK4) and thus halting the cell from progressing from the G1 phase to the S phase. Its production is promoted by Rb and E2F being tightly linked. However in the presence on E7, which displaces E2F, this creates an accumulation of E2F and thus an over expression of inactive P16, the high quantity of P16 can be used as a marker for the diagnosis of cervical cancer.

HPV types and diseases

There are over 100 types of HPV types, not all cause any symptoms in humans, here is a list of the most common symptoms associated with HPV and the type which causes said disease:

• Common warts: 2 and 7
• Plantar warts: 1, 2 and 4
• Flat cutaneous warts: 3 and 10
• Angogenital warts: 6, 11, 42, 43, 44, 55 and more
• Genital malignancies: 16, 18, 31, 33, 35, 39, 45, 51
• Epidermodysplasia verruciforms: 5 and 8 especially, however there are 15+ types.
• Focal epithelial hyperplasia (oral): 13 and 32
• Oral papillomas: 6, 7, 11, 16 and 32

HPV genes and their roles

• Control region: LCR
• Promoter: p97
• Oncogenes: E5, E6 and E7
• Viral replication: E1
• Viral transcription factor: E2
• Capsid formation: L1 and L2
• Viral maturation: E4

Location prevalence

Areas in which HIV are quite prominent, cervical cancer is also. This is because HIV causes those infected to become immunocompromised, thus they are significantly more susceptible to the HPV virus. Also, western countries have more modernised screening programmes, and also the use of contraception, such as condoms, is more widely used, thus incidence would be decreased. 

Risk factors

These include:

• Adolescence/young age
• Early onset of sexual activity (at, or before the age of 17, hence the HPV vaccine given to young girls)
• Number of sexual partners (e.g greater than 5)
• Risk of partners (they could be very sexually active and be of a "high" risk)
• Oral contraceptives
• Cigarette smoking (cotinine, a chemical found in tobacco, can be found within the cervical mucus, and has the ability to damage DNA)
• Lack of screening (those who've had pap smears every 3 years have 1/10 the risk of someone who hasn't had any)
• Immunosuppression

Likelihood

An approximate of 80% of the sexually active population will be involved with the HPV infection. It is very likely most people will come into contact with HPV at some point in their lives. 3/4 Americans between the ages of 15-50 have been infected by HPV at some point, however, the immune system can usually clear it. It is the most common sexually transmitted diseases. 

HPV Prevalence 

HPV can be present in a woman's cervix and vulva, inside the surrounding areas of the vagina and anus. It is common to be asymptomless. It can be obtained and then cleared without the recipient ever knowing. It is most commonly passed on unintentionally from those without symptoms, hence why it is so common. 

HPV persistence

Most women who are aware of having contracted HPV clear it, however it is unknown how many clear it completely, as it can go into remission after an active period, this could be for months or even years. Sometimes it is not entirely eliminated from the body. Infections can re-emerge from latency when said person becomes in a state of temporarily or permanent immunodeficiency  related to sunburn, pregnancy, stress both physical and emotional, cancer chemotherapy and HIV infection. It is impossible to state how long a person has been infected or when it will emerge. 

HPV and males

Men express HPV infection in the form of penile, perennial and urethral papillomas. They are much more likely to become a HPV carrier, and they will not develop cancer, as it is rare for papillomas to manifest themselves onto oncogenic cells. In order to swab for a pap analysis, a urethral swab can be taken to collect exfoliated cells however this is seldom practised. There is no FDA approved test for men thus clinically it is hard to be tested.Plus, unless there are symptoms such as aPl wart, from non-oncogenic HPV types such as type 6 and 11, men are less likely to attend any screenings.

Papanicolaou (PAP) staining

This is a classic stain, it involves the following:

• Haematoxylin stain: this is used to stain cellular nuclei, a blue - black colour.
• OG-6 counterstain: OG = Orange 6 and 6 refers to the conc. used of phosphotungstic acid, may have other variants 5 and 8. This stain colours keratin yellow.
• Second EA counterstain: EA = Eosin Azure

If performed appropriately, the stained specimen will show hues of the whole colour spectrum, red, orange, yellow, green, blue and violet. If the specimen was well prepared before the stain, the cellular nuclei will be a strong blue - black colour. Cells which have a high keratin content will be stained yellow, glycogen will also stain yellow. Superficial cells will be a orange - pink colour, and intermediate and parabasal cells will be a green/blue turquoise colour. Metaplastic cells will stain both green and pink. 

Under the microscope, it will be possible to establish whether the cells are CIN 1, 2 or 3. As the grade of the lesion increases (1-3) the cytoplasm to nucleus ratio will decrease, this is because the cells are becoming less differentiated.

Liquid based cytology

The instrument is inserted into the cervix and rotated 360 degrees in order to get cell samples from all parts, endocervix, transformation zone and the ectocervix, it is then taken out and placed into a fluid, which is then centrifuged and examined. It is them smeared out and a pap stain is used upon the sample, this provides a clean and clearer appearance. The material may last for up to 2 months and many more slides can be created from it (20-30). An automatic robot can screen these samples. Other smear tests pose problems with brushes which can damage the cells, and also with cells being left on the brushes. 

Koilocytes

These are cells which have a halo around their nuclei, this can be witnessed with a PAP smear. These cells are squamous epithelial cells which have undergone a transformation due to the infection of HPV, thus they are a positive indictor of the presence of HPV.

Smear results

If the results of a smear return back as "normal" this means there were no nuclear abnormalities  and the patient will be placed on a routine recall. 

If the results of a smear return back as "inadequate" this means there was insufficient material present, or it was poorly spread/fixed. The vision of the cells could be obscured by debris. This occurs in approximately 9% of all smears. The patient must repeat the smear, if this occurs more than 3 times consecutively, the patient must be referred for a colposcopy.

If the results of a smear return back as "border line dyskaryosis" this means that there was nuclear changes present which are not normal, however it cannot be certain whether the changes definitely represent dyskaryosis. This occurs in 5-10% of all smears. This means that the patient should repeat the smear in 6 months, as in a general case most smears will have reverted back to norma. After 3 consecutive normal smears, the patient is placed back on normal recall. If the abnormality persists after 3 smears, or gets worse, the patient is referred to a colposcopy. If in a 10 year period there are 3 borderline or results become more severe, the patient should be referred to a colposcopy. 

If the results of a smear return back as "mild dyskaryosis" this means that nuclear abnormalities are indicative of low grade CIN. The patient must then repeat the smear in 6 months. If after 3 more, results return back to normal, the patient is placed on normal recall. If the changes persist on 2 occasions  the patient is referred for a colposcopy. If in a 10 year period patient has 3 moderate results, they're referred to a colposcopy. If CIN1 is histologically confirmed, the management options are not entirely clear, depending on the results it could be watched or it could be treated.

If the results of a smear return back as "moderate dyskaryosis' this means that nuclear abnormalities reflect a probable case of CIN2, this occurs in approximately 1% of all smears. The patient will be referred to a colposcopy. If the CIN stage is histologically confirmed, treatment is provided to remove areas of abnormal cells. 

If the results of a smear return back as "severe dyskaryosis" or worse, this means that nuclear abnormalities are reflecting probable CIN3. This occurs in approximately 0.6% of smears. The patient should be referred to a colposcopy or in some rare cases, make an urgent referral to a gynaecological oncologist if invasive carcinoma is suspected. If the CIN3 stage is histologically confirmed, treatment is provided to remove areas of abnormal cells. 

NHS screening

The Nation Health Service Cervical Screening Programme (NHSCSP) is regarded as one of the best in the world, it has around 80% attendance and carries out 4 million smears a year. As a result, the number of deaths from cervical cancer in the UK and Wales is below 1000 every year. Statically due to an increase in sexual activity and a decrease in age, there should be around 5000 deaths. They offer smear tests at the age of 25 and every 3 years after that until 50 years, and from then on every 5 years. Annual tests are performed for 2 years after CIN 1 treatment, and 10 years after CIN 2 or 3 treatment. 

VILI

Squamous epithelium contains glycogen, however precancerous lesions and invasive cancer contains little to no glycogen. Iodine is glycophilic and is taken up by the squamous epithelium, staining it a mahogany brown/black colour (VILI negative). Columnar epithelium does not contain glycogen. Immature metaplasia and inflammatory lesions are at most, only partially glycogenated and when stained, it appears as scattered, ill-defined take up areas. Precancerous lesions and invasive cancer do not take up iodine, and will appear as well-defined thick mustard/saffron yellow areas (VILI positive). 

Cervical biopsies

There are two ways of sampling the the cervix, via the use of a punch biopsy, where the instrument takes small areas leaving small holes around the cervix, or a cone biopsy, where a scalpel cuts out the middle area of the cervix. 

Staging

These stages are provided by histological and clinical assessment e.g. MRI scan.

Prognosis

In accordance to the stages, here is the % survival rate of patients:

• Stage I: 91.5%
• Stage IIa: 83.5%
• Stage IIb: 66.5%
• Stage IIIa: 45%
• Stage IIIb: 36%
• Stage IV: 14%

Treatment

Vaccinations

Prophylactic vaccines are virus like particles, they consist of just the recombinant L1 structural protein, which has the ability of self assembly it contains no DNA, but the capsid itself can initiate an immune response, as it resembles intact viruses. They're non-infectious,  and very immunogenic. 

If women are vaccinated against HPB types 16, 18, 31, 33, and 45 before exposure, there will be an 85% reduction in cancer risk, decrease of 45-70% in the frequency of abnormal pap smears, a decline in abnormal pap smears within months, and a reduction in the incidence of cervical cancer would take a decade. 

However, it is important to remember, that not all cervical cancers are caused by HPV!