Cyto pathology

Cytopathology

Is the science of interpretation of cells obtained from human body by natural or atrifecial means.

Advantages of cytopathology

1.Easy,safe ,Quick,painless and repeatable.2.It doesn’t require hospitalisation or anasthesia.3.Sometime it has theraputic application.4.It can provides materials for culture and sensitivity tests.5.It can provides materials for immunohistochemistry.6.It may indicate the histogenesis of tumour

Disadvantages of cytopathology

1.It lacks the microanatomical and architectural changes. 2.Its accuracy is sometime less than histopathology .

Applications of cytopathology

1.Diagnosis of malignant and pre malignant lesions. 2.Follow up of treated cases for recurrence. 3.Treatment of haematoma or cysts. 4.Diagnosis of some infections . 5.Diagnosis of hormonal status in vaginal smears.

Complications of cytopathology

It is free from serious complications.The main complications include: 1.Heamatoma. 2.Infection. 3.Bleeding. 4.Anaphylactic shock 5.Spreed of tumor cells. 6.Pneumothorax in FNAC from lung.

Cytopathological features of malignancy

1.Features in the nuclei 2.Features in the cytoplasm

Features of malignancy in the nuclei

1.Size 2.Shape 3.Characters of chromatin 4.Mitosis

Features of malignancy in the cytoplasm

1.size relative to the nuclei i. e N/C ratio 2.shape 3.staining properties 4.phagocytosis

Requirements for good cytopathology report

1.Good clinical history 2.collection of adequate sample 3.proper processing ( fixation, staining ) 4.Good cytopathologist

Gynaecological cytopathology

1.Anatomy and histology of female G.T.:

Collection of specimens

1.Vaginal pool aspirate 2.Smear from lateral vaginal wall 3.Cervical scrape 4.Endocervical smear 5.Endometrial smear

Indications of each types of smears

1.Vaginal pool aspirate: Detection of carcinoma of FGT, specially endometrial and vaginal carcinoma 2.Smear from lateral Vaginal wall:for Hormonal assesment 3.Cervical Scrape: Carcinoma of cervix 4.Endocervical smear:for endocervical Ca. 5.Endometrial smear: For endometrial Ca.


How to collect the sample
1. Avoid vaginal douch and sexual intercourse 24 hours before smear 2. Best time around mid cycle, but it can be taken at any time 3.smear should be collected before bimanual examination 4.speculum should be unlubricated 5.Both vag. & Cx smears should be taken and fixed while they are wet in 95% alcohol

Clinical history

Name,Age, date of cycle, presenting symptom,(bleeding discharge, pelvic pain) clinical diagnosis, history of surgery, radiotherapy,hormones,IUCD, cytotoxic drugs

Cytopathology report

1.Describe the smear 2.diagnose 3.give further advise

Normal Smear from FGT

Normal constituent of smear from FGT depend on age , date of the cycle and site of the smear

Normal contents of Vaginal and cervical smears

1. Epithelial cells: a. squamous epithelium b. columnar epithelium c. endometrial cells 2. Nonepithelial cells: a.WBCs b. RBCs c. others like spermatozoa

Squamous Epithelial cells

1.Superfecial squames: 2.Intermediat squames: 3.Parabasal cells: 4.Basal cells :

Endocervical cells

They are tall columnar cells with basal nucli and cilliated margion.A vacule of mucus may be seen in the cytoplasm. A thread like extention indicate its attachment to basment membrane Cytologically they look columnar when veiwed from side & honny comb appearance when veiwed from top

Endocervical cells

Endometrial cells
1.Endometrial glandular cells 2.Endometrial stromal cells

Nonepithelial cells

1.Leukocytes 2.Spermatozoa

Satisfactory smear

1.Satisfactory vaginal smear 2.Satisfactory cervical smear

Causes of Inflammatory changes in cervical and vaginal smears

1.Bacterial: e.g.staph, G.C.,E-coli etc 2.Viral: Herpes. HPV etc 3.Fungal:Candida, Leptothrix etc 4.Protozoal:Trichomonas vaginalis(T.V.)

Features of inflammation in smear

1.Increase in number of polymorphs 2.Degenerative and regenerative changes 3.Dirty background 4.The cause can sometime be seen e.g. T.V.Monilial hyphae & spores

Bacterial infection

G.C., staphylococi, E-coli streptococi etc. The cytological picture is that of nonspecific inflammatory smear. No specific feature could be seen

Viral infection

Herpes simplex type II and human papilloma viruses Type 6, 11,16,18,33,&35 are the most common viral infection. The importance of HPV is its carcinogenic effect Cytologically: nuclear enlargment,multinucleation with characteristic molding of the nuclei Inclusion bodies may be seen

Trichomonas vaginalis

Result from infection by trichomonas vaginalis Cytologically: Inflammatory smear Eosinophilia and Perinuclear hallo of the squamous epithelial cells . Presence of trichomonas vaginalis parasite

Fungal infection

1.Monilial infection;Infection by candida albicans.The fungus has thin septated hyphe with spores Affect immunocompromised patients Cytologically : Inflammatory smear contain fungal hyphe and spores

Fungal infection

Leptothrix: Thick nonseptated fungi, commonly seen in association with T.V. Cytologically: Inflammatory smear contain the fungi and sometime T.V.

Other infections

Actinomycosis: infection by actinomycetes. Commonly associated with copper IUCD Cytologically: Inflammatory smear, contain radiating coloni of actinomycetes

Chlamydia

Strictly intracellular organism Cytologically : inflammatory smear , with intracellular presence of trophozoid or cysts

Gardnerella vaginalis

Gardrenella vaginalis is gm variable coccobacilli commonly associated with anerobic bacterial infection Cytologically: Inflammatory smear contain clue cells.(clue cell is a squamous epithelium covered by the M.O.)

cytolysis

This term is used to describe fragmentation of the cytoplasm by lactobacilli while the nuclei remain intact . Cytolysis may affect endocervical cells, intermediat and parabasal cells. The denuded nuclei can be recognised for malignant changes

Squamous metaplasia

Transformation of columnar epithelium to squamous epithelium.in the smear they apear rounded or polyhedral about the size of parabasal cells,they have orange cytolasm, vesicular nuclei and tinny nucleolus. Mature squamous metaplasia are similar to intermediet cells,while Immature squamous metaplasia are similar to parabasal cells

Metaplasia (cont)

Atypical squamous metpalasia shedded in loose clumps , have eosinophilic cytoplasm and hyperchromatic nuclei

Koilocytotic atypia

In viral infection squamous cells show vaculisation of the cytoplasm .The nuclei show atypical changes in form of hyperchromasia, presence of inclusion bodies


In the cervix, koilocytotic change with human papillomavirus infection, is seen here, with vacuolization of epithelial cells.

Cervical erosion

A clinical term used to describe extention of endocervical epithelium outside the endocervical canal. Cytologically it show inflammatory smear with numerous sheets of hyperplastic endocervical cells and metaplastic squames with a lot of fresh blood.

leukoplakia

A cinical term used to describe whitish discolouration of any mucus membrane. Its importance is that it may harbour dyslasia or carcinoma in situ. cytologicaly the smear contain numerous anuclear squames.

Benign cellular changes (hyperkeratosis), x200

Evaluation of hormonal status
Estrogen produces rapid and complete maturation. The pattern of exfoliation is in single cells so the smear contain superfecial mature squames in single pattern. Abscence of estrogen produces complete atrophy of the squamous epithelium i.e the smear is atrophic &contain mainly parabasal cells

Evaluation of hormonal status(cont)

Progesteron produces rapid proliferation with production of intermediat cells pattern .more important is the pattern of exfoliation , it produces desquamation of clumps of folded intermediet cells

Evaluation of hormonal status (cont)

When both estrogen and progesteron are combined as in luteal phase,progesteron antagonise the effects of estrogenand result in absence of superfecial cells due to massive desequamation. Androgen produce intermediate cell pattern and the shedding in single cells and small group of cells.

Smear in menstrual phase

Blood, endometrial cells,WBCs and intermediat cells.

Proliferative phase

After menstruation the smear pattern is that of intermediat cell pattern few endometrial cells may be seen for few days after menstruation. Superfecial squames the start to appear and increase in number until the smear becomes entirely superfecial squames in single cell distribution ( Ovulatory smear)

Luteal phase

After ovulation there is decrease in number of superfecial cells,The pattern become intermediat cell pattern shedded in clumps Few days befor menstruation endometrial cells apeare in the smear and WBC increases in number

At menopause

The smear is mainly parabasal cells i.e atrophic smear Atrophic smear also seen in postnatal and in lactating smears



In pregnancy
1.There is loss of cyclical changes in the smear 2.Intermediat cells arrangesd in clumps with folding of their cytoplasm .The nuclei are elongated (Cigar shape) and a yellow colour or orange colour appear arround the nucleus(Due to high content of glycogen) These cells are called Navicular cells or Oyster cells

Factors affect hormonal assessment

1.Inflammatory process. It look more mature. 2.Cytolysis 3.Drugs: e.g. hormones digoxins 4.Other treatments like cautery,Radiotherapy

Methods of hormonal assessment(Indices of squamous epithelium)

1.Karyopyknotic index (K.I.) 2. Acidophylic index using schorr stain.(A.I.) 3. Maturation index (M.I.)

Malignant and premalignant lesions of cervix

Etiology of cervical cancer: It is multifactorial and include environmental,chemical,genetic,viral e.g.HPVetc. It is abscent in vergins More common in unstable &multiple marriges Early exposure to sex Penile factors

Pathogenesis of cervical cancer

Malignant transformation occure when the epithelial cells are actively undergoing squamous metaplasia in adolescence and early pregnancy. The process of carcinogenesis involve two mechanisms Intiation and Promotion.

Pathogenesis of Cx cancer (cont)

Intiation: Carcinogen gain access to susceptible active metaplastic cells and alter them Promotion: occure by repeated exposure to intiator or or new nonspecific agents which increase normal and abnormal mitotic activity resulting in development of clone of cell which overcome immunological mechanism of the body and result in cancer


Morphological evolution of squamus cell carcinoma of Cx
Reserve cell hyperplasia-squamous metaplasia--dysplasia-carcinoma in situ-- invasive cancerThis sequence may progress or regress at any time

Dysplasia

Failure of epithelial cells to mature adequatly .According to the severity it may be: Mild Moderate Sever Cells shedded from dysplastic epithelium is called dyskaryotic cells.


At low magnification, note the demarcation between the normal cervical mucosa at the left and the dysplastic epithelium at the right.

At higher magnification, cervical dysplasia is demonstrated. Note the disorderly development of the cells from the basement membrane upward at the right, compared to normal maturation at the left.

Dyskaryotic cells

A cell with abnormal nucleus (showing some features of malignancy) but with relatively normal cytoplasm . According to the severity it may be: Mild dyskaryotic cells Moderate dyskaryotic cells Sever dyskaryotic cells

Dyskaryotic cells (cont)

These types depend on subjective assessment depending on degree of nuclear and cytoplasmic changes. Dyskaryotic cells from deeper layers ( i.e intermediet or parabasal layers) are more significant than dyskaryotic cells from superfecial layers.

Cytological picture of dysplasia

The severity of the lesion is reflected on the smear. Mild dysplasia: Superfecial dyskaryotic squames,nuclear abnormality is not so sever . Usually associated with inflammatory smear

Cytological picture of dysplasia (cont)

Moderate dysplasia: Superfecial and intermediet dyskaryotic squames. Sever dysplasia: Dyskaryotic parabasal cells together with superfecial and intermediet dyskaryoic cells.


Cytologic features of dysplasia are seen on Pap smear. Some of the cells at the center show increased nuclear/cytoplasmic ratio, with darker and more irregular nuclei than the normal squamous cells with large amount of cytoplasm and small, pyknotic nuclei

Carcinoma in situ of cervix

Alteration of surface epithelium that histologically resmble cervical cancer but there is no invasion

Cytological picture of Carcinoma in situ

Mixture of dyskaryotic cells and cancer cells with variable differentiation from anaplastic cells to well differentiated keratinised cells. Clean background

Microinvasive cancer

Mean very early but unquestionable invasion in the form of sharp thin tongue of epithelium invade the underlying tissue. Sometime extention of the epithelium is enmass in the stroma

Invasive carcinoma of cervix

Any cancer extend beyond the anatomical bounderies of the surface epithelium or endocervical glands is regarded as invasive cancer.It may be: Squamous cell carcinoma 90-95% Adenocarcinoma 5%

Fungating, exophytic tumor is seen on the cervix, typical for invasive squamous cell carcinoma


A biopsy reveals invasive squamous cell carcinoma of the cervix.

Cytological picture of inasive cancer of cervix

Malignant cells with many bezar cells Dyskaryotic cells from the margion of the lesion. Dirty background containing necrotic debries and blood.

Malignant cells

A cell with abnormal nucleus and abnormal cytoplasm E.g. Tadpole cells and fiber cells

Squamous cell carcinoma, x200.

The cytologic features of invasive squamous cell carcinoma are demonstrated on a Pap smear. Note the cells with hyperchromatic, pleomorphic nuclei and increased nuclear/cytoplasmic ratio. The background has inflammatory cells

Causes of false negative smears

LGSIL (mild dysplasia and human papillomavirus-associatedchanges), x400. Marked air-drying artifact and heavyinflammation obscure cellular detail in this smear; inparticular, chromatin texture is difficult to evaluate. Nuclearirregularity suggests a preneoplastic lesion. A careful searchturned up unequivocal LGSIL in a better-preserved portion ofthe smear.



Recurrent carcinoma of cervix
Any carcinoma regardless of the method of treatment should be followed by cytological examination to detect any recurrence , as ca of cervix may recure or it may have multifocal origion

Effects of radiation

Nuclear ballooning , multinucleation & vaculation and breakdown of nuclei. Cytoplasmic vaculation

Natural history of dysplasia & carcinoma in situ

Dysplasia & Ca in situ are precursor of invasive ca. They may regress by treatment of infection and taking smear. They may progress from mild dysplasia to sever dysplasia ca in situ and invasive ca 30-60 % of ca in situ progress to invasive ca.It may take 10-20 Y to progress.

Pap grading

Grade I Normal Grade II Inflammatory Grade III Marked atypia .No enough evidence for diagnosis of cancer. Grade IV Ca in situ Grade V Invasive ca

Dysplasia of endocervical epithelium

Abnormal arrangment of cells, together with the features of dyskaryotic cells (i.e hyperchromasia, coarse chromatin pattern, pleomrphism etc)

Carcinoma of endocervix

It is usually adenocarcinoma Cytologically : The smear contain sheets of malignant glandular cells i.e. Large vesicular nuclei , prominant nucleoli, together with all the other features of malignancy. The cytoplasm contain mucus vacules indicating its glandular origion.


Endometrial carcinoma
Endometrial carcinoma is also adenocarcinoma. Vagival smear is the usefull smear for detection of endometrial ca. Cytologically the smear contain sheets of malignant glandular cells .usually in small clusters .theis nuclei are small rounded with coarse chromatin pattern

Endometrial stromal sarcoma and other malignantt lesion e.g. G.T.

Rare lesions . Cytologically: Stromal sarcoma consist of sheets of malignant spindle cells with haemorrhagic background. Gestational tumour present as large pleomorphic and bezar cells and haemorrhagic background

Screening for cervical cancer

Cervical cancer is a potentially fatal cancer It can be prevented if diagnosed at the stage of dysplasia or carcinoma in situ. Cervical cytopathology is the ideal method for detection of these lesion at an early stage. Screening for cx cancer usually start at the age of 18 Y or at the begining of sexual activity.Cx & Vg. Smears are taken every 1-3 years

Rational for cytological examination of the cervix

Carcinoma of cervix usually develop from precursor dysplasia or ca in situ. These lesion are symptomless and produce no visible signs. So the only practical methods of detecting these lesion is by cytological examination.At these stages the disease is curable .i.e. You can cure potentially lethal disease.

Cytopathology of Respiratory System

Anatomy and Histology
The respiratory system start from nose , nasopharynx, larynx, trachea, bronchi, bronchial branches and alveoli. Histologically: The R.S is lined by pseudostratified columnar cilliated epithelim, part of the larynx lined by squamous epithelium,the brochi contain goblet cells in between the cilliated columnar cells.The alveoli lined by pneumocytes and contain alveolar macrophages

Samples from respiratory system

1.Sputum:it should be deep cough specimen 2.Brochial wash.:Through bronchoscope 3.Bronchial brush:Using bronchial brush 4.Percutaneous aspiration.

Preparation of smear

Sputum sample: Smears are prepared from freshly received sputum without fixatives.blood stained sputum should be selected.Using forceps, sputum is smeared on labeled slides and fixed in 95% alcohol before drying. Bronchial wash: Received in trap plastic bag, centerfuge and prepare smear from the sediment and fix in 95% alcohol

Preparation of smear (cont)

Bronchial brush: smears are prepared by the endoscopist from the plastic brush and fixed in 95% alcohol.

Normal contents of smears

Smears from Upper Respiratory Tract: Superfecial and intermediet squamous cells, few polymorphs, columnar cells.It does not contain alveolar macrophages. Smears from lower Respiratory tract: Inaddition to the above contents in contain alveolar macrophages which is an indication of adequate specimen

 Normal bronchial epithelium, x200

Inflammatory smear

Squamous cell carcinoma in sputum

Adenocarcinoma, x400

Body fluid cytology
Body fluid include: Pleural fluid Ascitic fluid Pericardial fluid CSF Joint fluid

Normal cells of body fluid

Mesothelial cells: Rounded or oval cells scattered singly and in small group Contain small vesicular nuclei sometime binucleated or multinucleated, Basophilic cytoplasm with small cytoplasmic projections may be seen Few histiocytes and occasional lymphocytes

Inflammatory conditions

The volum is increased . A collectable amount is considered as abnormal . Cytologically it depends on the type of inflammatory condition. In pyogenic infections polymorphs are the predominate cells together with mesothelial cells and few lymphocytes In chronic inflammatory condition lymphocytes are predominate e.g.T.B.

Malignant effusion

In primary mesothelioma: Rare presents as clumps, balls of cells with all the features of malignancy . In metastatic lesion, the type of cells depends on the primary origion e.g. adenocarcinoma, oat cell carcinoma , melanoma etc In CSF the main indication is involvment by lymphoma and leukaemia

fine needle aspiration

Lymph node, fine needle aspiration


Metastatic melanoma, x200

GastroIntestinal Tract

The sites accessible by endoscopy to obtain materials for cytological examination include: 1.Esophagus 2.Stomach 3.Duodenum up to ampulla of vater and ERCP to common bile duct. 4.Rectum, colon up to caecum

Normal contents

Esophagus and stomach: 1.Squamous epithelium 2.Gastric mucosal cells 3.Few inflammatory cells 4.Swallowed alveolar and bronchial cells.

Normal content:

Colon: 1. colonic mucosal cells 2.Few inflammatory cells

Advantages of GIT cytology

1. Collect materials from wider area than biopsy. 2.Detection of early gastric cancer 3. Good adjunct to biospy 4. Easy, safe ,repeatable,ecconomic,quick

Methods of obtaining samples

1. Through gastroscope by brush& wash. 2. Gastric lavage

Lesions of GIT diagnosable by cytopathology

1. Malignant lesion of esophagus;e.g. Squamous cell carcinom adenocarcinoma. Lymphoma. 2. Nonmalignant lesions e.g. Gastric ulcer,monilial infection of esophagus e.c.

Squamous cell carcinoma of Esophagus

Cytologically a well keratinised malignant squames are seen in a necrotic background

Adenocarcinomas of esophagus

Cytologically sheets of malignant glandular cells with its vesicular nuclei , prominant nucleoli and some times the cytolasm contain mucus vacules. The cells are forming loose sheets and scattered singly.

Lymphoma

Cytologically large number of monomorphic lymphoid cells usually larger than small lymphocytes, contain nucleoli .The cells are scattered singly.

Gastric ulcer

Cytologically :Necrotic background, large number of inflammatory cells ,sheets of bland looking gastric mucosal cells, the cells are of similar size and shape, form honycomb appearance. Some of the sheets show hyperplastic changes

The urinary system

Application of cytopathology in U.T.: The main applications of cytopathology in urinary system is detection of malignancy. Nonmalignant lesions include infections parasitic ova etc Diagnosis of malignancy is mainly applicable to lower urinary tracy (Bladder) Lesions high up is difficult to diagnose due to degerative changes during the journey of the cells downto the bladder.

Samples from urinary tract

Urine the commonest sample used, rarely brush and wash materials may be collected. Freshly passed(voided) urine is centrifuged and smears are prepaired from the sedement and fixed in alcohol.Then stained by H&E or Pap stain. Incatheterised patient urine should be collected from the catheter not from bag)

Normal contents of UT

Mainly urothelial cells a rounded or oval cells with one or more bland looking vesicular nuclei and basophilic cytplasm.scattered singly and in small groups. Few inflammatory cells may be found. Squamus epithelium from urethra In catheterised urine the number of sheets are numerous due to the irritation by the catheter.

Carcinoma of the bladder

Most cancer of bladder are transitional cell carcinoma Sometime squamous metaplasia may be associated. Adenocarcinoma is rare arising from remnant of uracus

Cytopathology of UT

Transitional cell carcinoma present as malignant urothelial cells, in loose sheets and and papillary clusters, with large hyperchromatic cells .These changes depends on the grade of the tumour .The background is bloody and contain necrotic materials. Suamous differentiation is represented by presence of squamus or squamoid cells.

Adenocarcinoma of Bladder

Rare cytologically like any other adenocarcinoma

Malignant lesion in kidneys

The diagnosis of Renal cell carcinoma has been reported but practically it is difficult.



Avantages of Cytology in UT
1. Provide preoperative diagnosis 2. Good adjunct to biopsy 3.Easy, saf repeatable,ecconomic. Accurate (40-100%) 4.Detection of recurrence is one of the most important application of cytology 5. Screening for early detection in high risk group.

Fine Needle Aspiration Cytology (FNAC)

It is the technique of obtaining cells and tissue fragments from organ that do not shed cells spontaneously for cytopathological examination.e.g. Brast mas, L.N. Liver etc by the use of disposable synge and needle.

Technique

A prelabled slides, fixative,are prepaired.The site of aspiration is cleaned by alcohol.The technique is performed by holding the mass by the fingers of one hand and the needle is passed in the lesion by the other hand. A suction is applied and while negative pressure is applied the needle is passed in and out in the lesion in different direction. The suction is released, and the needle is withdrawn, the aspirated materials are smeared on the slides, fixed and stained

FNA technique

FNA technique

FNAC technique

Advantages of FNAC
1.Easy, safe, repeatable, relatively painless,ecconomic, accurate. 2.May have theraputic application. 3.Provides materials for culture and sensitivity, AFB etc 4. May diagnose the histogenesis of the lesion

Disadvantages

Limited accuracy specially in some lymphomas and endocrine organs. May foul the patient and delay treatment

Complications of FNAC

Almost nil Haematoma Inection Anaphylactic shock in hydatid cyst aspiratrion Pneumothorax in lung aspiration

Hepatocellular carcinoma, Diff-Quik, x200

Pleomorphic adenoma, x200

Apocrine metaplasia

Fat necrosis

Duct carcinoma

Lobular carcinoma