Saturday, 31 August 2019

Immune tolerance at endometrium and association with Rec preg losses.

It is said that endometrial NK cell act as traffic control- by preventing entry of friendly classical T cells). We are already ware that “Endometrial Leukocyte population –as separate from peripheral blood CD 56+ cells   activated by IL 15 , There are  three subtypes  end level .These age 1) u-NKcells 2) Normal macrophages and third type of 3) uterine leukocytes present are T calls.

 Of all NK cells present in endometrium are comprise of CD 56, CD 16 but in periphery it is CD 15 that potentiates the actions of CD 56.
One has to remind that 90  & of endometrial u-N cells are comprise of CD 56, CD 16. As mentioned earlier presence of high no of u-NKcells may initiate aborting again. HLA-G & amount of interferon-¥ plays a significant role in the development process of spiral arteries as occurs in preg.

Dr Pal’s long list: - My policy & what I do:-My policy is to draw blood one time and to minimise the visits to pathology Lab by the couple.  It has been concluded that psycho-immunology is an important factor. The members may rightly asked list are quite costly and even if a deficiency is identified one  is very certain whether such cause is playing the role of abortion or else if a drug is available . For instance if Cong Protein-C deficiency or Protein S deficiency will treatment be effective? Who will supervise the following preg if treatment  is implemented? 

What are routine list of investigations that have to be asked as a protocol of Lab assessment as preconception medical fitness? Complete Heamogram, H [LC, Group, & typing, Serology for STD –HIV, TPHA in particular, Viral screening, Thyroid screening, PRL, Hepatitis serology, Rubella screening, Glycaemic  profile, LFT. Renal profile (at least Creatinine), Routine Pelvic USG. Urine RE



& in myoma:  - Antigen screening mechanism by T cells
 Immunity is of two types. .
A)  B-cell immunity: - or humoral (antibodies mediated like –immunoglobulin’s of varying types) –noncellular  i.e.   Serum immunity.
B)  T cell immunity: - this is a Cell mediated immunity and is expressed by Macrophages, Lymphocytes, Lymph nodes and   & effector  T Cells. Unlike B cell immunity T cells are not only restricted in blood stream and present in fair number in tissues as well. In control of diseases both B and T cell immunity are required. Cell culture technique can now differentiate detail functioning / dynamics of T cell immunity mechanism.

C)  CLASSIFICATION OF T CELLS Mechanisms:-

D) Cytotoxic  T cells

 a) Cytotoxic T cells: - These are also called   effector T cells, Carrying CD 8 markeràlysing the host cells which was invaded by the pathogenic organism/ foreign antigens. B) T helper Cell this carry CD 4 marker):-: This type of cells in conjunction with
Macrophages /phagocytes destroy the invading enemy by releasing chemical agents called CYTOKINES. Therefore in short T cells can destroy foreign antigens either by lysis (cytotoxic CD8 cells) or can destroy by production id cytokines (by CD4 Cells).

What are cytokines?

Therefore chemical agents are (cytokines are secreted by B-Helper cells to release
 Such bioactive substances.
T (Cellular) & B (humoral) cell Dysfunction
Disorders of Immune dynamics can cause lot of diseases and even may be prime cause extinction of human race in the centuries to come. It is for this system we are still alive and their help is most needed in all most all diseases including natural reproduction.
Types of B cell Dysfunction: - a) Over reaction b) Immunodeficiency (reduced defense reaction) c) Autoimmunity-incorrect / inappropriate response/reaction.
MHC:
MHC:- Major Histocompatibility Complex:- These are  one type of friendly receptors present on the surface of T cells-by which it recognize its own inherent  antigens and do not response from reacting or producing cytokines. But when foreign antigens present to the T cell surface then T cell start producing antibodies.
.T cell response is of two types: - a) Cytotoxic response b) Helper function:
CD8+ T cells (have power to recognize MHC class I antigens) but CD4+ T cells have power to recognize MHC class II antigens.
Antigen screening mechanism by T cells is an inborn power whether to act or not to react. To whom to attack, when to attack, how to attack the offending invader (in this case may be paternal antigen/antigens) buy with some dignity so that endometrium itself is not damaged or menorrhagia/Metrorrhagia  do not ensue as continued effect of sperm antigens/ placental antigens/antigen.
Memory cells.    Are capable of producing messenger cytokines after recognize fresh entry of foreign antigens.  Memory cells.    Are capable of producing messenger cytokine molecules. This can in turn attract and promote production of effector  T cells.  
:The issue of  efficacy of progesterone in idiopathic Rec abortion:-However ,  Efficacy of progesterone in apparently  idiopathic Rec abortion as raised: May I inform my dear members that  there is am trial going on "PROMISE"-WWW,medscinet.net/promise at UK & Netherland. This is also applicable in cases of Sub chorionic haematoma.
To what extent the exhaustive investigations will help the couple? Idiopathic  in about 40-50% cases of RSA !!!:-Clinicians feel sorry and feel ashamed (at least I do  feel guilty & full of   shyness prevail in my face)  :- if no cause is identified  after exhaustive Lab tests &  investigations  spending  so  much  money for investigations:-In many studies time and again , it has been firmly concluded, that in about 40-50% cases of RSA-the etiology remain uncertain even in best centers including  centers of western part of globe( unlike our resource poor country) . What are routine list of investigations that have to be asked as a protocol of Lab assessment as preconception medical fitness? Complete Heamogram, H [LC, Group, & typing, Serology for STD –HIV, TPHA in particular, Viral screening, Thyroid screening, PRL, Hepatitis serology, Rubella screening, Glycaemic profile, LFT. Renal profile (at least Creatinine), Routine Pelvic USG. Urine RE


What is the problem in investigating cases of RPL who can afford? Multiple subtle ( for which no marker is available till date –say genes/enzymatic deficiency at endometrial level/Interleukins and Cytokines /Knell population/Factors which takes overcharges from  CL of preg to placental factors for continuation pregnancy): taken together may eventually lead to RA.

Will detailed ,special & costly investigations be helpful  in  treating subsequent pregnancies? Is it not true that some minor other diseases or deficiencies will remain unnoticed &unrecognized and she will any case abort as that diseases is not tretaed: _Will investigation help the clinician? Is to what extent? Multiple subtle factors taken together may eventually lead to RA. Such researches and many senior clinicians have also pointed out that multiple subtle factors taken together may eventually lead to RA. They (researches) have stressed that a number of factors may be operating in many cases and to make the matter more worse for the   clinician’s investigators have opined that in different pregnancies different factors may be may be operatoionable.  That is their belief. That is why possibly TLC has a major role in the management of such syndrome/ disease
To treat or not to treat if a defect is observed in Lab tests –Will that TR be effective at all?? It is couples & doctor’s choice as there is no guarantee that TR is going to offer guarantee about a Live birth: The inhibition of prescribing septic drugs stems from Int recommendations as Level of evidence!! Doctors are hesitant if level of evidence is 4!! :-Should we regard/disregard a Lab report and whether TR o such defect will materially help the distressed woman? What is meant by Int recommendations as LEVEL of Evidence? Treat a How difficult for us (clinicians) to lay importance on a   particular disease as the prime cause of abortion if Lab report so suggest: - Recommendations issued by different International Agencies have be to be followed by us??   Many agencies have stratified the efficacy of different diseases as a causative factor RSA  as level of evidence in the format of evidence level-+1 ,2 ++, 3,  4 as in many other diseases. It is more relevant so far as RCOG guidelines are followed. If recommendation is like “evidence level 3 or say 4-then one is hesitant to prescribe drug for such presumed diseases in cases of RPL as evidence is, say to say, lacking firmly.

Stratification of etilogy-oriented investigations, thereby to cut short list of investigations & revise the plan the list of investigations (to cut short long list of investigations as per gest age of RSA):  What was the gest age when previous 2-3 abortions occurred-stratification of etilogy-oriented investigations,  The very philosophy of stratification of etilogy-oriented investigations”- If one believes in this proposition :-Then what special tests to be followed”thogh  as a clinician, most of us perform following tests for Rec spont abortion cases ,particularly where there have been more than 3 abortions and couple and their relatives are prepared to  pay any amount of money before planning for third pregnancy
Different  attitudes of different clinicians  about list of investigates :-Many clinicians have different  attitudes about list of investigates and some astute clinicians stratify the list according to gestational age of abortions –a) WHWTER the previous losses were Early Looses(  then they strongly  considers :- a) endocrine factor & b)chromosomal/ c) genetic causes d) uterine natural killer cells-uNK cell population—a very common cause of idiopathic early abortion .possibly more than genetic or chromosomal disorders) ANF, tests for aPL ab
. But if Losses occurred later after 10 week the possibility of following abnormalities be cone stronger. E g.  1) Structural malformations of uterus,-best by hysteroscopy) & at the same time Lap to exclude minimal endometriosis which cannot be picked by USG or say asymptomatic but continue to release oocyte toxic materials which eventually affect “ovum pick up mechanism”. This release of toxic materials and gameto-toxic cytokines is however also true for Latent Kochs which can be occasionally be picked by peritoneal sampling. Hypercoagulability disorders-Acqd or congenital,   / platelate COUNT BY DIFF METHODS,PLATELATE FUNCTIN IF POSSIBLE, PT,PTTK Hhcy, environmental diseases, smoking,   addiction, stress and possibly immunological & above all psychological as the “week of previous risk” approaches, Hystero-Laparoscopy,
If PREVIOUS abortions occurred after say 8weeks-it is presumable that though  many clinicians don’t accept this stratification of etilogy-oriented investigations, Instead they(clinicians) refer the couple concerned asking for “Recurrent Abortion Panel”:. I don’ know whether such practice is appropriate or not. What are the member’s advice & their practice pattern??

.

Detailed List of investigation:-Pros & Cons: - - To investigate at one time (one stop Investigatios particularly who have more than three pregnancy losses excluding those Lab tests which have already done earlier. Admittedly such list is long (most of the members, I am sure will disagree with me about my philosophy of getting all tests done) may cost Rs 25,000/- to Rs 35,000/- according to quality and name & fame of Lab List is, though exhaustive but excludes usual preconceptionally tests are
 1) Autoantibodies-ANA, Anti-ds-DNA
 2) Tests for A-APL ab 3) Thrombophilia screening3) 4) HSG /3-D usg/SIS choice /preference of Gynecologist concerned.4) LPD as a precursor of early preg failure-by causing poor endometrial development. ?? 5)Infection screening:-Chlamydial & Gonococci screening if not done earlier, &Mycoplasma & Listerosis are not available at Kolkata. 6)  Homocysteine Normal value of plasma  is 6-14 µ.mol/L, Mostly, low Homo signify defective Vit B12 metabolism due possibly to Foliate or other enzymatic disorders(inborn), Hyper level of Homo causes much damage not only in the  preg outcome but abnormal accumulation of homocysteine in serum (-H.Hys ) is a marker/ warning  lab parameter of thrombo-embolism    including Coronary   thrombosis,7) Estimate AMH & Gaunt AFC: _Low AMH, AFC & DOR:-as a cause of poor oocyte qualityà failure to development of onwards growth of embryo//foetus (disordered post zygotic events)-genetic error by aged Oocyte .akin to trisomy in aged females.
8) Detailed & special sperm function tests”
9) Parental karyotyping:







 Scrum progesterone in day 22 of unstimulated cycle in cases of early failures-admittedly questionable because most of us suppl P after UPT is +ve, but if P value in luteal phase is suboptimal (if < 3-4 ng/ml)-but fortunately such P level is above 10 ng/ml in mid/late luteal pages then there is possibly no indication for LPD support in the cycles when they will be attempting for pregnancy. The date of drawing blood in irregular unstimulated cycle may be done either as LH Urine test /FM .then we have to suppl after,

-   6) Infection screening:-Chlamydial & Gonococci screening if not done earlier, &Mycoplasma & Listerosis are not available at Kolkata. &


8) Homocysteine Normal value of plasma  is 6-14 µ.mol/L, Mostly, low Homo signify defective Vit B12 metabolism due possibly to Foliate or other enzymatic disorders(inborn), Hyper level of Homo causes much damage not only in the  preg outcome but abnormal accumulation of homocysteine in serum (-H.Hys ) is a marker/ warning  lab parameter of thrombo-embolism    including Coronary   thrombosis,
9)  Estimate AMH & Gaunt AFC: _Low AMH, AFC & DOR:-as a cause of poor oocyte qualityà failure to development of onwards growth of embryo//foetus (disordered post zygotic events)-genetic error by aged Oocyte .akin to trisomy in aged females.
10) Detailed & special sperm function tests”
11) Parental karyotyping:

12) - Elevated APAs are equally prevalent among women experiencing unexplained infertility, recurrent implantation failure, and recurrent pregnancy loss. Heparin and aspirin are successful in the treatment of elevated APA among women with recurrent miscarriage but not with recurrent implantation failure.IVIg has been successful in the treatment of recurrentmiscarriage and recurrent implantation failure among women with elevated APA and/or NK cell activity Intralipid is effective in the treatment of women experiencing reproductive failure who display elevated NK cell activity. For immunological factors, other than APA ARGUABLY, there is no clear-cut benefit of any intervention...
Should we routinely sample endo for uNK cells during hysteroscopy? If so will be beneficial, and if so to what extent? What does excessive u-u-NKcells population in endometrium signify?? Is it a danger signal of again another recurrent abortion? Few points about u-NK cell population as assayed by immunohistchemistry –sampling done at the time of hysteroscopy along with Kochs diagnosis by stain, special  culture, exclusion of synechiae, polyp or septum s other commonly observes diseases in diagnosed at hysteroscopy? It is said that endometrial NK cell act as TRAFFIC CONTROL- by preventing entry of friendly classical T cells). We are already ware tat “Endometrial Leukocyte population –as separate from peripheral blood CD 56+ cells   activated by IL 15) have three subtypes I end level .These age 1) u-NKcells 2) Normal macrophages and third type of uterine leukocytes present are T calls, Of all NK cells present in endometrium are comprise of CD 56, CD 16 but in periphery it is CD 15 that potentiates tie cactions of CD 56.
One has to remind that 90& of endometrial u-N cells are comprise of CD 56, CD 16, AS mentined earlier presence of high no of u-NKcells may initiate abortin again. HLA-G & amount of interferon-¥ plays a significant role in the development process of spiral arteries as occurs in preg.

Dr Pal’s long list: - My policy & what I do:-My policy is to draw blood one time and to minimise the visits to pathology Lab by the couple. 2 It has been concluded that psycho-immunology is an important factor.
 The members may rightly asked list are quite costly and even if a deficiency is identified one  is very certain whether such cause is playing the role of abortion or else if a drug is available . For instance if Cong Protein-C deficiency or Protein S deficiency will treatment be effective? Who will supervise the following preg if TR is implemented? 

What are routine list of investigations that have to be asked as a protocol of Lab assessment as preconception medical fitness? Complete Heamogram, H [LC, Group, & typing, Serology for STD –HIV, TPHA in particular, Viral screening, Thyroid screening, PRL, Hepatitis serology, Rubella screening, Glycaemic profile, LFT. Renal profile (at least Creatinine), Routine Pelvic USG. Urine RE



& in myoma:  - Antigen screening mechanism by T cells
 Immunity is of two types. .
E)   B-cell immunity: - or humoral (antibodies mediated like –immunoglobulin’s of varying types) –noncellular i.e.   Serum immunity.
F)   T cell immunity: - this is a Cell mediated immunity and is expressed by Macrophages, Lymphocytes, Lymph nodes and   & effector T Cells. Unlike B cell immunity T cells are not only restricted in blood stream and present in fair number in tissues as well. In control of diseases both B and T cell immunity are required. Cell culture technique can now differentiate detail functioning / dynamics of T cell immunity mechanism.

G) CLASSIFICATION OF T CELLS Mechanisms:-

H) Cytotoxic  T cells

 a) Cytotoxic T cells: - These are also called   effector T cells, Carrying CD 8 markeràlysing the host cells which was invaded by the pathogenic organism/ foreign antigens. B) T helper Cell this carry CD 4 marker):-: This type of cells in conjunction with
Macrophages /phagocytes destroy the invading enemy by releasing chemical agents called CYTOKINES. Therefore in short T cells can destroy foreign antigens either by lysis (cytotoxic CD8 cells) or can destroy by production id cytokines (by CD4 Cells).

What are cytokines?

Therefore chemical agents are (cytokines are secreted by B-Helper cells to release
 Such bioactive substances.
T (Cellular) & B (humoral) cell Dysfunction
Disorders of Immune dynamics can cause lot of diseases and even may be prime cause extinction of human race in the centuries to come. It is for this system we are still alive and their help is most needed in all most all diseases including natural reproduction.
Types of B cell Dysfunction: - a) Over reaction b) Immunodeficiency (reduced defense reaction) c) Autoimmunity-incorrect / inappropriate response/reaction.
MHC:
MHC:- Major Histocompatibility Complex:- These are  one type of friendly receptors present on the surface of T cells-by which it recognize its own inherent  antigens and do not response from reacting or producing cytokines. But when foreign antigens present to the T cell surface then T cell start producing antibodies.
.T cell response is of two types: - a) Cytotoxic response b) Helper function:
CD8+ T cells (have power to recognize MHC class I antigens) but CD4+ T cells have power to recognize MHC class II antigens.
Antigen screening mechanism by T cells is an inborn power whether to act or not to react. To whom to attack, when to attack, how to attack the offending invader (in this case may be paternal antigen/antigens) buy with some dignity so that endometrium itself is not damaged or menorrhagia/Metrorrhagia do not ensue as continued effect of sperm antigens/ placental antigens/antigen.
Memory cells.    Are capable of producing messenger cytokines after recognize fresh entry of foreign antigens.  Memory cells.    Are capable of producing messenger cytokine molecules. This can in turn attract and promote production of effector T cells.  

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Kindly give suggestions regarding Recurrent first trimester loss (blighted ovum, missed abortion) evaluated found to have elevated homocystein
methylcobamin 1500 b.d ,folic acid, Arginine
 How high is it? What is pts age? Any other factor for RPL
 History: - What about her nutrition, any F/H/O of Koch’s?, Explore family tree for genetic diseases, Life style disease /habit of either partn
  Common Tests”- In addition suggestions made by some forum members, may carry out Complete haemogram, Thalassaemia screening, Viral screening, Metabolic screen (HBA1C & PPBS) & endocrine screen( TSH, PRL, Total testosterone).

Thrombophilias Screening, A...
 Uncommon Tests:- AMA, ANA. Hystero-Laparoscopy-Minimal endometriosis & septal diseases of uterus.I often perform this test. To me this operation should be in the list of common tests, but many differ due to cost / invasiveness.
Psycho-immunology in the treatment of RSA is an emerging subject. Support of the family members & relatives/ friends .But in the days of consumers many doctors are hesitant to assure the couple “All is well, Next time such unfortunate event will not occur. I am with you. God is going to bless you this time.”.
 Summary:-Above all, uterine malformations, endocrine disorders and advanced age of any one partner are the usual causes of RSA.
May read RCOG green top guideline no. 17 (issued on April, 2011). In about 50% cases the etiology remains unexplored. But in such situation, I do prescribe, albeit imperially a course of antibiotics for pelvic . Infection like doxycycline & Azithromycin.
 One of my close friends presented with similar history,
elevated homocysteine with recurrent losses.
later on on further evaluation due to affordability,
she was found to have mutation in MTHR,
currently on nutritional supplements,L-methyl folate the active form of frolic acid apart from b12 etc.other

 hereditary thrombophilia screen were negative like factor V Leidin.. mutation in MTHR.
 MTHFR gene mutation is common in Asians and doesn't require treatment! That cannot be cause for RPL
 Thank u mam smile emoticon yeah her treating RPL : Q What % of conception proceed to  live birch?? Ans: Human reproduction is remarkably inefficient; nearly 70% of human conceptions do not survive to live birth.

What is the commonest cause of spont abortion?? Ans;- the most common cause for spontaneous loss, is  fetal aneuploidy is particularly in the first trimester of pregnancy. Although losses may be due to A) owing to de novo fetal aneuploidy occur at similar frequencies among women with sporadic and recurrent losses, B) Equal no have  couples additional associated genetic factors and C) The other causes of RSL /RPL are  monogenetic etiologies.

Q, How useful is Genetic testing of the products of conception from couples experiencing two or more losses?? Ans:-Genetic testing of the products of conception from couples experiencing two or more losses may aid in defining the underlying etiology and in counseling patients about prognosis in a subsequent pregnancy.

How important is Parental karyotyping of couples  Ans:-Parental karyotyping of couples who have experienced recurrent pregnancy loss (RPL) will detect some couples with an increased likelihood of recurrent fetal aneuploidy; this may direct interventions. Parental karyotyping is better than on POCs, only because 70% SABs have chromosomal abnormalities, but that does not change the outcome for future pregnancies. On the other hand, if either of the parents are carriers of balanced translocation [6% possibility], then the recurrence risk will depend on the specific translocation

How important is PGS?? The utility of Perimplantation genetic analysis in couples with RPL is unproven, but new approaches to this testing show great promise.
Haemoglobin nature by HPLC, Blood Group, & typing, Serology for STD –HIV, TPHA in particular, Viral screening, Thyroid screening (T .profiler) , PRL*, Hepatitis serology, Rubella screening, Glycemic profile, LFT. Renal profile (at least Creatinine), Routine Pelvic USG-any myoma, endometriosis, adenomyosis,  Urine RE ,Karyotypes, Day 3LH*, .TPHA, Thrombophilia screening . Homocysteine*, Ant-do DNA ab, Anti smooth ms ab, Anti thymoglobulin ab, ANA, anti-microsomal ab, ANCA(anti neurtrophil cytoplasmic ab,  USG serum testis , Insulin to exclude PCO, Hysteroscopy, HSG, GTT , Chlamydia, Brucella screening , Cervical swab & culture,. Mycoplasma, HSV, 3-D USG, Polyspermia, Serum Progesterone Tets for SLE (like a) anti cardiolipin an GPL U/ml & Lupus anticoagulant ab like divvy (LA 1) & APTT (PTT-LA)  Tets four cong hypercoagulable diseases like deficient if protein C, S & Ante thrombin III


What is the problem in investigating cases of RPL who can afford? Multiple subtle ( for which no marker is available till date –say genes/enzymatic deficiency at endometrial level/Interleukins and Cytokines /Knell population/Factors which takes overcharges from  CL of preg to placental factors for continuation pregnancy): taken together may eventually lead to RA.

Will detailed ,special & costly investigations be helpful  in  treating subsequent pregnancies? Is it not true that some minor other diseases or deficiencies will remain unnoticed &unrecognized and she will any case abort as that diseases is not treated: _Will investigation help the clinician? Is to what extent? Multiple subtle factors taken together may eventually lead to RA. Such researches and many senior clinicians have also pointed out that multiple subtle factors taken together may eventually lead to RA. They (researches) have stressed that a number of factors may be operating in many cases and to make the matter more worse for the   clinician’s investigators have opined that in different pregnancies different factors may be may be operatoionable.  That is their belief. That is why possibly TLC has a major role in the management of such syndrome/ disease
To treat or not to treat if a defect is observed in Lab tests –Will that TR be effective at all?? It is couples & doctor’s choice as there is no guarantee that TR is going to offer guarantee about a Live birth: The inhibition of prescribing septic drugs stems from Int recommendations as Level of evidence!! Doctors are hesitant if level of evidence is 4!! :-Should we regard/disregard a Lab report and whether TR o such defect will materially help the distressed woman? What is meant by Int recommendations as LEVEL of Evidence? Treat a How difficult for us (clinicians) to lay importance on a   particular disease as the prime cause of abortion if Lab report so suggest: - Recommendations issued by different International Agencies have be to be followed by us??   Many agencies have stratified the efficacy of different diseases as a causative factor RSA  as level of evidence in the format of evidence level-+1 ,2 ++, 3,  4 as in many other diseases. It is more relevant so far as RCOG guidelines are followed. If recommendation is like “evidence level 3 or say 4-then one is hesitant to prescribe drug for such presumed diseases in cases of RPL as evidence is, say to say, lacking firmly.

Stratification of etilogy-oriented investigations, thereby to cut short list of investigations & revise the plan the list of investigations (to cut short long list of investigations as per gest age of RSA):  What was the gest age when previous 2-3 abortions occurred-stratification of etilogy-oriented investigations,  The very philosophy of stratification of etilogy-oriented investigations”- If one believes in this proposition :-Then what special tests to be followed”thogh  as a clinician, most of us perform following tests for Rec spont abortion cases ,particularly where there have been more than 3 abortions and couple and their relatives are prepared to  pay any amount of money before planning for third pregnancy
Different  attitudes of different clinicians  about list of investigates :-Many clinicians have different  attitudes about list of investigates and some astute clinicians stratify the list according to gestational age of abortions –a) WHWTER the previous losses were Early Looses(  then they strongly  considers :- a) endocrine factor & b)chromosomal/ c) genetic causes d) uterine natural killer cells-uNK cell population—a very common cause of idiopathic early abortion .possibly more than genetic or chromosomal disorders) ANF, tests for aPL ab
. But if Losses occurred later after 10 week the possibility of following abnormalities be cone stronger. E g.  1) Structural malformations of uterus,-best by hysteroscopy) & at the same time Lap to exclude minimal endometriosis which cannot be picked by USG or say asymptomatic but continue to release oocyte toxic materials which eventually affect “ovum pick up mechanism”. This release of toxic materials and gameto-toxic cytokines is however also true for Latent Kochs which can be occasionally be picked by peritoneal sampling. Hypercoagulability disorders-Acqd or congenital,   / platelate COUNT BY DIFF METHODS,PLATELATE FUNCTIN IF POSSIBLE, PT,PTTK Hhcy, environmental diseases, smoking,   addiction, stress and possibly immunological & above all psychological as the “week of previous risk” approaches, Hystero-Laparoscopy,
If PREVIOUS abortions occurred after say 8weeks-it is presumable that though  many clinicians don’t accept this stratification of etilogy-oriented investigations, Instead they(clinicians) refer the couple concerned asking for “Recurrent Abortion Panel”:. I don’ know whether such practice is appropriate or not. What are the member’s advice & their practice pattern??

.

Detailed List of investigation:-Pros & Cons: - - To investigate at one time (one stop Investigatios particularly who have more than three pregnancy losses excluding those Lab tests which have already done earlier. Admittedly such list is long (most of the members, I am sure will disagree with me about my philosophy of getting all tests done) may cost Rs 25,000/- to Rs 35,000/- according to quality and name & fame of Lab List is, though exhaustive but excludes usual preconception ally tests are
 1) Autoantibodies-ANA, Anti-do-DNA
 2) Tests for A-APL ab 3) Thrombophilia screening3) 4) HSG /3-D us/SIS choice /preference of Gynecologist concerned.4) LPD as a precursor of early preg failure-by causing poor endometrial development. ?? 5)Infection screening:-Chlamydial & Gonococci screening if not done earlier, &Mycoplasma & Listerosis are not available at Kolkata. 6)  Homocysteine Normal value of plasma  is 6-14 µ.mol/L, Mostly, low Homo signify defective Vit B12 metabolism due possibly to Foliate or other enzymatic disorders(inborn), Hyper level of Homo causes much damage not only in the  preg outcome but abnormal accumulation of homocysteine in serum (-H.Hys ) is a marker/ warning  lab parameter of thrombo-embolism    including Coronary   thrombosis,7) Estimate AMH & Gaunt AFC: _Low AMH, AFC & DOR:-as a cause of poor oocyte qualityà failure to development of onwards growth of embryo//foetus (disordered post zygotic events)-genetic error by aged Oocyte .akin to trisomy in aged females.
8) Detailed & special sperm function tests”
9) Parental karyotyping:







 Scrum progesterone in day 22 of unstimulated cycle in cases of early failures-admittedly questionable because most of us suppl P after UPT is +ve, but if P value in luteal phase is suboptimal (if < 3-4 ng/ml)-but fortunately such P level is above 10 ng/ml in mid/late luteal pages then there is possibly no indication for LPD support in the cycles when they will be attempting for pregnancy. The date of drawing blood in irregular unstimulated cycle may be done either as LH Urine test /FM .then we have to suppl after,

-   6) Infection screening:-Chlamydial & Gonococci screening if not done earlier, &Mycoplasma & Listerosis are not available at Kolkata. &


8) Homocysteine Normal value of plasma  is 6-14 µ.mol/L, Mostly, low Homo signify defective Vit B12 metabolism due possibly to Foliate or other enzymatic disorders(inborn), Hyper level of Homo causes much damage not only in the  preg outcome but abnormal accumulation of homocysteine in serum (-H.Hys ) is a marker/ warning  lab parameter of thrombo-embolism    including Coronary   thrombosis,
9)  Estimate AMH & Gaunt AFC: _Low AMH, AFC & DOR:-as a cause of poor oocyte qualityà failure to development of onwards growth of embryo//foetus (disordered post zygotic events)-genetic error by aged Oocyte .akin to trisomy in aged females.
10) Detailed & special sperm function tests”
11) Parental karyotyping:

12) - Elevated APAs are equally prevalent among women experiencing unexplained infertility, recurrent implantation failure, and recurrent pregnancy loss. Heparin and aspirin are successful in the treatment of elevated APA among women with recurrent miscarriage but not with recurrent implantation failure.IVIg has been successful in the treatment of recurrentmiscarriage and recurrent implantation failure among women with elevated APA and/or NK cell activity Intralipid is effective in the treatment of women experiencing reproductive failure who display elevated NK cell activity. For immunological factors, other than APA ARGUABLY, there is no clear-cut benefit of any intervention...
Should we routinely sample endo for uNK cells during hysteroscopy? If so will be beneficial, and if so to what extent? What does excessive u-u-NKcells population in endometrium signify?? Is it a danger signal of again another recurrent abortion? Few points about u-NK cell population as assayed by immunohistchemistry –sampling done at the time of hysteroscopy along with Kochs diagnosis by stain, special  culture, exclusion of synechiae, polyp or septum s other commonly observes diseases in diagnosed at hysteroscopy? It is said that endometrial NK cell act as TRAFFIC CONTROL- by preventing entry of friendly classical T cells). We are already ware tat “Endometrial Leukocyte population –as separate from peripheral blood CD 56+ cells   activated by IL 15) have three subtypes I end level .These age 1) u-NKcells 2) Normal macrophages and third type of uterine leukocytes present are T calls, Of all NK cells present in endometrium are comprise of CD 56, CD 16 but in periphery it is CD 15 that potentiates tie cactions of CD 56.
One has to remind that 90& of endometrial u-N cells are comprise of CD 56, CD 16, AS mentined earlier presence of high no of u-NKcells may initiate abortin again. HLA-G & amount of interferon-¥ plays a significant role in the development process of spiral arteries as occurs in preg.

Dr Pal’s long list: - My policy & what I do:-My policy is to draw blood one time and to minimise the visits to pathology Lab by the couple. 2 It has been concluded that psycho-immunology is an important factor.
 The members may rightly asked list are quite costly and even if a deficiency is identified one  is very certain whether such cause is playing the role of abortion or else if a drug is available . For instance if Cong Protein-C deficiency or Protein S deficiency will treatment

What tests for Recurrent Preg Losses?? What investigations?? Rationality?? A thread bare analysis


There are about 40 known tests in a case of RPL albeit most are of theoretical benefit as in many such diseases there is no known  therapy. As such,  there are many experienced clinicians who stratify cases according the time of abortion, if at all such miscarriages occur at almost similar periods of gestations and try to limit the number of tests.  There are equally another group of clinicians who assume one or two types of cause based on his/ her clinical experience and initially try to exclude only those particular diseases, This type of clinical thought is often termed by the clinicians as etilogy-oriented investigations. There are yet another group of clinicians who don’t like to miss any diagnosis and they (clinicians) refer the couple concerned to a reputed Lab asking for “Recurrent Abortion Panel” and also to seek advise hematologist for any Thrombophilic or other hematologic diosrder including hypercoagulable starts which we gynaecologist don’t understand so well. That is not unfair and I should say that if affordable is a good practice patter.  

However, the following tests are very commonly performed in most cases in nonpreg cases even after one pregnancy loss .Admittedly , these are often   routine tests and most are  unrelated to preg loss . These are   1) Haemoglobin nature by HPLC, 2) Blood Group, & typing, Indirect Coombs tests  3) Serology for STD –HIV, TPHA in particular, 4) Viral screening including hepatitis serology 5) Thyroid screening (Thyroid profile) , 6) PRL*, Serum Progesterone & Day: 3 LH* , 7) Rubella screening, 8) serum tests PP Sugar & PP insulin to exclude  insulin resistance , OGTT i.e. Glycemic profile, 9) LFT.10)  Renal profile (at least creatinine), 11) Routine Pelvic USG-any myoma, endometriosis, adenomyosis,  12) Urine RE , 13) USG:-AFC & other imaging information  as available 14) HSG  15) Diag lap-Hysteroscopy .16) AMH :: However those  marked with * are for cases of vary  early abortions . Of note in most of such disease some form tr can be offered to improve fertility potential .Therefore these are first line investigations (modifiable factors). Readers may add or subtract any of tests depending on his/her belief and accumulated experience, as above mentioned list is from my experience and not approved by any academic bodies.

Part II
1)             Thrombophilia screening.  2)  Tests for SLE like a) anti cardiolipin an GPL U/ml & Lupus anticoagulant ab like divvy (LA 1) & APTT (PTT-LA)  3) Tests four cong hypercoagulable diseases like deficient if protein C, S & Anti thrombin III
 4) Ant-do DNA ab, 5)  Anti smooth ms ab, 6) Anti thymoglobulin ab, 7) ANA, 8) anti-microsomal ab, 9) ANCA(anti neurtrophil cytoplasmic ab,  10) Karyotypes 11) Hysteroscopy/ HSG, 12) , Chlamydia, 13) Brucella screening , 14)  Cervical swab & culture,. 15) Mycoplasma, 16) HSV, 17) 3-D USG, 18) Homocysteine*,  19) Polyspermia,

What is the problem in investigating cases of RPL who can afford? Ans: If no therapy is available then there is little benefit of testing. . There are multiple subtle diseases/ deficiencies  ( known / as yet unknown)  for which no marker is available till date –say genes/enzymatic deficiency at endometrial level. Similarly abnormalities of  different interleukins and cytokines /Knell population/Factors which takes over charges from  CL of preg to placenta  for continuation of smooth progress of pregnancy) are difficult to diagnose but some agents are used like  antioxidants, I V immunoglobulins.: Taken together the lack of beneficial or blastocyst friendly growth factors may eventually lead to RA. Empirical Tr are 1) antioxidants 2) life style changes 3) empirical tr with antibiotics for presumed / established reproductive tract infections .  4) I V immunoglobulins :-Ivy has been successful in the treatment of recurrent  miscarriage and recurrent implantation failure among women with elevated APA and/or NK cell activity 5) Intralipid is effective in the treatment of women experiencing reproductive failure who display elevated NK cell activity. For immunological factors, other than APA arguably, there is no clear-cut benefit of any intervention.AS such question aeries
should we routinely sample endo for uNK cells (uterine natural killer cells ) during hysteroscopy? If so will it be  beneficial to concerned woman , and if so to what extent? What does excessive u-u-NKcells population in endometrium signify??

Are we missing something in the investigation panel ?? Will detailed special & costly investigations be helpful in treating subsequent pregnancies? Is it not true that some minor other diseases or deficiencies for which no tests are  available will remain unnoticed & unrecognized. In such situations and she will any case abort as such subtle diseases / syndromes were never suspected and therefore possibility of such diseases will remain untreated
Will investigation help the clinician? If yes,  to what extent? Multiple subtle factors taken together may eventually lead to RA. Such researches and many senior clinicians have also pointed out that multiple subtle factors taken together may eventually lead to RA. They (researches) have stressed that a number of factors may be operating in many cases and to make the matter more worse for the   clinician’s investigators have opined that in different pregnancies different factors may be may be operatoionable.  That is their belief. That is why possibly TLC has a major role in the management of such syndrome/ disease
To treat or not to treat if a defect is observed in Lab tests –Will that Tr be effective at all?? It is couples & doctor’s choice as there is no guarantee that Tr is going to offer about a live birth: The inhibition of prescribing such  drugs stems from Int recommendations as Level of evidence!! Doctors are hesitant if level of evidence is 4!! :-Should we regard/disregard a Lab report and whether Tr of such defect will materially help the distressed woman? What is meant by Int recommendations as LEVEL of Evidence? Now we can understand  how difficult is for us (clinicians) to lay importance on a   particular disease as the prime cause of abortion if Lab report so suggest: - Recommendations issued by different International Agencies have be to be followed by us,    Many agencies have stratified the efficacy of different diseases as a causative factor RSA  as level of evidence in the format of evidence level-+1 ,2 ++, 3,  4 as in many other diseases. It is more relevant so far as RCOG guidelines are followed. If recommendation is like “evidence level 3 or say 4-then one is hesitant to prescribe drug for such presumed diseases in cases of RPL as evidence is, say to say, lacking firmly.
Stratification of etilogy-oriented investigations, thereby to cut short list of investigations & revise the plan the list of investigations (to cut short long list of investigations as per gest age of RSA):  What was the gest age when previous 2-3 abortions occurred-stratification of etilogy-oriented investigations,  The very philosophy of stratification of etilogy-oriented investigations”- If one believes in this proposition :-Then what special tests to be followed though  as a clinician, most of us perform following tests for Rec spont abortion cases ,particularly where there have been more than 3 abortions and couple and their relatives are prepared to  pay any amount of money before planning for third pregnancy
Different  attitudes of different clinicians  about list of investigates :-Many clinicians have different  attitudes about list of investigates and some astute clinicians stratify the list according to gestational age of abortions –a) whether the previous losses were Early Looses(  then they strongly  considers :- a) endocrine factor & b)chromosomal/ c) genetic causes d) uterine natural killer cells-uNK cell population—a very common cause of idiopathic early abortion .possibly more than genetic or chromosomal disorders) ANF, tests for aPL ab
. But if Losses occurred later after 10 week the possibility of following abnormalities be cone stronger. e g.  1) Structural malformations of uterus,-best by hysteroscopy) & at the same time Lap to exclude minimal endometriosis which cannot be picked by USG or say asymptomatic but continue to release oocyte toxic materials which eventually affect “ovum pick up mechanism”. This release of toxic materials and gameto-toxic cytokines is however also true for latent Kochs which can be occasionally be picked by peritoneal sampling.
Hypercoagulability disorders-acqd or congenital,   / platelate count by diff methods,  platelate function if possible, PT, PTTK Hhcy, environmental diseases, smoking,   addiction, stress and possibly immunological & above all psychological as the “week of previous risk” approaches, Hystero-Laparoscopy,
If previous abortions occurred after say 8 weeks-it is presumable that though  many clinicians don’t accept this stratification of etilogy-oriented investigations, Instead they(clinicians) refer the couple concerned asking for “Recurrent Abortion Panel”:. I don’ know whether such practice is appropriate or not. What are the member’s advice & their practice pattern??

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Detailed List of investigation:-Pros & Cons: - - To investigate at one time (one stop Investigatios particularly who have more than three pregnancy losses excluding those Lab tests which have already done earlier. Admittedly such list is long (most of the members, I am sure will disagree with me about my philosophy of getting all tests done) may cost Rs 25,000/- to Rs 35,000/- according to quality and name & fame of Lab List is, though exhaustive but excludes usual preconception ally tests are
 1) Autoantibodies-ANA, Anti-do-DNA
 2) Tests for A-APL ab 3) Thrombophilia screening3) 4) HSG /3-D us/SIS choice /preference of Gynecologist concerned.4) LPD as a precursor of early preg failure-by causing poor endometrial development. ?? 5)Infection screening:-Chlamydial & Gonococci screening if not done earlier, &Mycoplasma & Listerosis are not available at Kolkata. 6)  Homocysteine Normal value of plasma  is 6-14 µ.mol/L, Mostly, low Homo signify defective Vit B12 metabolism due possibly to Foliate or other enzymatic disorders(inborn), Hyper level of Homo causes much damage not only in the  preg outcome but abnormal accumulation of homocysteine in serum (-H.Hys ) is a marker/ warning  lab parameter of thrombo-embolism    including Coronary   thrombosis,7) Estimate AMH & Gaunt AFC: _Low AMH, AFC & DOR:-as a cause of poor oocyte qualityà failure to development of onwards growth of embryo//foetus (disordered post zygotic events)-genetic error by aged Oocyte .akin to trisomy in aged females.
8) Detailed & special sperm function tests”
9) Parental karyotyping:







 Scrum progesterone in day 22 of unstimulated cycle in cases of early failures-admittedly questionable because most of us suppl P after UPT is +ve, but if P value in luteal phase is suboptimal (if < 3-4 ng/ml)-but fortunately such P level is above 10 ng/ml in mid/late luteal pages then there is possibly no indication for LPD support in the cycles when they will be attempting for pregnancy. The date of drawing blood in irregular unstimulated cycle may be done either as LH Urine test /FM .then we have to suppl after,

-   6) Infection screening:-Chlamydial & Gonococci screening if not done earlier, &Mycoplasma & Listerosis are not available at Kolkata. &


8) Homocysteine Normal value of plasma  is 6-14 µ.mol/L, Mostly, low Homo signify defective Vit B12 metabolism due possibly to Foliate or other enzymatic disorders(inborn), Hyper level of Homo causes much damage not only in the  preg outcome but abnormal accumulation of homocysteine in serum (-H.Hys ) is a marker/ warning  lab parameter of thrombo-embolism    including Coronary   thrombosis,
9)  Estimate AMH & Gaunt AFC: _Low AMH, AFC & DOR:-as a cause of poor oocyte qualityà failure to development of onwards growth of embryo//foetus (disordered post zygotic events)-genetic error by aged Oocyte .akin to trisomy in aged females.
10) Detailed & special sperm function tests”
11) Parental karyotyping:

12) - Elevated APAs are equally prevalent among women experiencing unexplained infertility, recurrent implantation failure, and recurrent pregnancy loss. Heparin and aspirin are successful in the treatment of elevated APA among women with recurrent miscarriage but not with recurrent implantation failure.IVIg has been successful in the treatment of recurrentmiscarriage and recurrent implantation failure among women with elevated APA and/or NK cell activity Intralipid is effective in the treatment of women experiencing reproductive failure who display elevated NK cell activity. For immunological factors, other than APA arguably, there is no clear-cut benefit of any intervention...
Should we routinely sample endo for uNK cells during hysteroscopy? If so will such test  be beneficial, and if so to what extent? What does excessive u-u-NKcells population in endometrium signify?? Is it a danger signal of again another recurrent abortion? Few points about u-NK cell population as assayed by immunohistchemistry –sampling done at the time of hysteroscopy along with Kochs diagnosis by stain, special  culture, exclusion of synechiae, polyp or septum s other commonly observes diseases in diagnosed at hysteroscopy? It is said that endometrial NK cell act as traffic control- by preventing entry of friendly classical T cells). We are already ware that “Endometrial Leukocyte population –as separate from peripheral blood CD 56+ cells   activated by IL 15 , There are  three subtypes  end level .These age 1) u-NKcells 2) Normal macrophages and third type of 3) uterine leukocytes present are T calls.