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NeoplasmDistinct metaplastic and inflammatory phenotypes in autoimmune and adenocarcinoma-associated chronic atrophic gastritis

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2023-01-01
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2017년 United European Gastroenterol J. 에 실린 내용입니다. Vanderbilt University School of Medicine, USA 의 Dr. Goldenring JR 의 연구팀에서 발표한 내용으로, Autoimmune gastritis (AIG)와 chronic atrophic gastritis (CAG)에서 유발된 adenocarcinoma 환자에서의 metaplasia, proliferation, immune cell populations의 차이를 본 연구입니다. 2022년 AIG와 Intestinal metaplasia를 정리하면서, 이러한 논문들이 바탕이 되고 계속 연구가 되었고 현재까지도 이어져 오면서 위염이나 위암과 같은 질환들에 대한 이해도가 높아질 수 있다고 생각해서 소개합니다. 

<Introduction>

 Helicobacter pylori (H. pylori)에 의한 chronic atrophic gastritis (CAG)는 이후 metaplastic lineage인 Spasmolytic polypeptide expressing metaplasia (SPEM) - 이제는 pyloric metaplasia or pseudo-pyloric metaplasia 라고 하는게 맞을 것 같습니다만 - 과 intestinal metaplasia 로 진행되고 결국 gastric adenocarcinoma의 neoplastic precusor 로 여겨지고 있습니다. Parietal cell loss는 chife cell의 SPEM으로의 trans-differentiation을 유도하게 되지만, DMP-777로 처리를 하면 dysplasi까지는 진행하지 않습니다. 하지만, inflammation으로 parietal cell이 손상이 되어 M2 macrophaes와 같은 추가적인 inflammatory process로 인해 proliferative metaplasia가 유도되면서 dysplasia나 cancer로 진행하게 됩니다. (Petersen CP, et al. Gastroenterology. 2014;146:1727-38.) Parietal cell loss (oxytic atrophy)는 adenocarcinoma과 높은 연관성을 가지고 있는 병리학적 소견이지만, AIG에서는 incidence가 낮은 수수께끼 (conundum) 에 대해 풀어보는 것이 이번 연구의 목적이 있다고 볼 수 있습니다.


<Materials and Methods>

CAG with adenocarcinoma: 12명은 endoscopic bx, 5명은 tissue specimens

AIG: with all autoimmune thyroiditis, endoscopy & serologic test (gastrin, chrmogranin, anti-parietal cell antibody)

H. pylori: histology (Giemsa and Warthin-Starry stainning), Intestinal metapalsia: type 1 (complete), type 2 (incomplete)

[Immunofluorescence stainning]

* Trefoil Factor 2 (TFF2): SPEM marker

* Mucin 2 (MUC2): Intestinal metaplasia marker

* CD68: Macrophage marker

* Myeloperoxidase (MPO): Neutrophil marker

* Conjugate-Griffonia simplicifolia II (GSII)-lectin: SPEM marker

* Counter stained with 40 ,6-diamidino-2-phenylindole (DAPI)

[Immunohistochemistry] 

* CD44 variant 9 (CD44v): SPEM marker

* DMBT1: Intestinal metaplasia marker (in human)

* Ki67: metaplastic mucosa marker

[Quantitation of immune cells and Ki67 staining] : Eosinophils (H&E stain, 10 HPF), Ki-67 staining quantitation


<Results>

[Clinical characteristics of the patient cohorts]

Table 1. Patient characteristics


Chronic atrophic gastritis group

Autoimmune atrophic gastritis group

Patient number

17

16

Age

65.9 ± 10.4

56.4 ± 10.6

Sex        Male

12

0

           Female

5

16

Mean serum gastrin

ND

913.4 ± 1195

Mean serum chromogranin A 

ND

336.2 ± 217.1

Neuroendocrine cell hyperplasia

ND

100%

Helicobacter pylori infection

62.5% (10/16)+

0%

Mean Eosinophil count/HPF (p=0.03)

3.50 ± 1.37

2.25 ±1.73

All quantitation is listed as mean ± SD / ND: Not determined. / +One patient had no information about H. pylori status.

<CAG with adenocarcinoma>

Patient demographics of chronic atrophic gastritis (CAG) group.

No

AGE

SEX

Cancer Pathology

Pathologic TNM Stage

Intestinal metaplasia subtype

H.pylori

Status

Mean Eosinophils

1

46

M

Poorly differentiated, signet ring cell adenocarcinoma

Not available

type I

negative

3

2

86

F

Well differentiated, mucinous adenocarcinoma

T4 G1 N1

type II

positive

4

3

48

M

Well differentiated, mucinous adenocarcinoma

Not available

type I

negative

0

4

79

M

Poorly differentiated, mucinous adenocarcinoma

T4 G3 N3

type I

negative

5

5

64

F

Poorly differentiated, diffuse adenocarcinoma

T4a G3 N2

type I

negative

5

6

75

M

Poorly differentiated, diffuse adenocarcinoma

T4a G3 N3

type I

ND

ND

7

63

M

Poorly differentiated, diffuse adenocarcinoma

T4a G3 N1

type 1

negative

3

8

65

F

 Poorly differentiated, diffuse adenocarcinoma

T4 G3 N3

type 1

positive

4

9

54

F

Poorly differentiated adenocarcinoma, intestinal type

T4a G3 N3b M1

type 1

positive

2

10

69

M

Intestinal type adenocarcinoma

T3 G1 N0

type 1

positive

4

11

64

M

Well-differentiated intestinal type adenocarcinoma

T4a G1 N3a

type 1

negative

2

12

58

M

Diffuse type antral adenocarcinoma

T4a G3 N3

type 1

positive

3

13

74

M

Intestinal type cardia adenocarcinoma

T3 G1 N1

type 1

positive

4

14

66

M

Intestinal type adenocarcinoma

T4 G1 N0

type 1

positive

3

15

68

M

Mixed type adenocarcinoma (predominantly diffuse)

T4a G3 N2

type 1

positive

4

16

68

F

Intestinal type adenocarcinoma

T3 G1 N0

type 1

positive

5

17

74

M

Intestinal type adenocarcinoma

Not available

type 1

positive

5


<Autoimmune gastritis>

Patient demographics of autoimmune atrophic gastritis (AIG) group.

No

Age

Sex

Serum Gastrin

Serum 

Chromogranin A

Serum 

Parietal cell Ab

Intestinal metaplasia subtype

neuroendocrine cell hyperplasia

Mean Eosinophil

1

47

F

>5000

102

yes

type I

yes

1

2

52

F

>1000

180

yes

type I

yes

1

3

61

F

1137

254

yes

type I

yes

5

4

72

F

35

169

yes

type I

yes

1

5

40

F

126.9

365

yes

type I

yes

3

6

78

F

218

ND

yes

ND

yes

5

7

51

F

654

420

yes

type I

yes

2

8

56

F

751

204

yes

type I

yes

2

9

46

F

44

66

yes

ND

yes

1

10

60

F

722

288

yes

type I

yes

1

11

66

F

345

238.6

yes

type I

yes

5

12

69

F

422

322

yes

type I

yes

3

13

51

F

749

820

yes

type I

yes

2

14

55

F

1041

664

yes

type I

yes

0

15

55

F

430

309

yes

ND

yes

4

16

44

F

1940

642

yes

ND 

yes

0


[Examination of metaplastic lineages in the atrophic stomach]

* All AIG patients: prominent parietal cell loss and TFF2-staining SPEM

* AIG patients: 75% (12/16) MUC2-positive goblet cell intestinal metaplasia (Figure 1(a)–(a3), 1(b)–(b3))

* All CAG: marked parietal cell loss and both TFF2-staining SPEM and MUC2-positive intestinal metaplasia (Figure 1(a)–(a3), 1(b)–(b3))

-> SPEM was more prominently represented in the atrophic stomach in AIG patients, intestinal metaplasia was present in both groups  

* CD44 variant (SPEM marker): Expression in SPEM (AIG & CAG patients), weak or absent Expression in intestinal metaplasia (AIG & CAG patients)  (Figure 1(c)–(c3)) 

* DMBT1 (Intestinal metaplasia marker): uniformly weaker in the AIG (Figure 1(d)–(d3))

* AIG: all type 1 (complete) metaplasia

* CAG: 16명/17명 type 1 (complete) metaplasia, 1명 type 2 (incomplete) metaplasia

Figure 1. . Metaplasia in the gastric mucosa of autoimmune gastritis (AIG) and chronic atrophic gastritis (CAG) patients. Top row: dual Alcian blue and periodic acid-Schiff (AB/PAS) staining of gastric mucosa in CAG ((a), (a1)) and AIG ((a2), (a3)) patients. Note AB staining of intestinal metaplasia in luminal cells, and PAS-staining of spasmolytic polypeptide expressing metaplasia (SPEM) at the bases of the gland. Second row: dual immunofluorescence staining of TFF2 (Trefoil Factor 2, SPEM marker, red) and MUC2 (Mucin 2, intestinal metaplasia marker, green) in gastric mucosa from CAG ((b), (b1)) and AIG ((b2), (b3)). In all cases nuclei were labeled with 40 ,6-diamidino2-phenylindole (DAPI) (blue). Images (a1), (a3), (b1) and (b3) are higher magnification as noted by white boxed outlines. Third row: immunohistochemical staining of CD44v (SPEM marker) in CAG ((c), (c1)) and AIG ((c2), (c3)) gastric mucosa. Bottom row: immunohistochemical staining of DMBT1 (Deleted in malignant brain tumors 1, intestinal metaplasia marker) for CAG ((d), (d1)), and AIG ((d2), (d3)) mucosa. Images (c1), (c3), (d1) and (d3) are higher magnification as noted by black-boxed outlines. Bar indicates 100 μm.

[Epithelial cell proliferation]

AIG와 CAG 모두 extensive metaplasia를 보였지만, AIG 환자의 mucosa의 높이가 낮아진 것으로 나타났습니다. 

AIG에서 Ki67-immunostaining cells 을 통해 살펴보면 metaplastic mucosal cells의 proliferation이 의미있게 낮아졌습니다. 이는, AIG에서 Metaplastic glands의 proliferationCAG보다 의미있게 저하되어 있다는 것을 의미합니다. (Figure 2)

Figure 2. Characteristics of proliferation in metaplastic mucosa. Representative immunostaining for Ki67 in (a) a chronic atrophic gastritis (CAG) patient and (b) an autoimmune gastritis (AIG) patient. (c) Quantitation of Ki67-positive cells demonstrates a markedly lower level of proliferation in AIG patients. In CAG patients, filled circles indicate females and open circles indicate males. *p = 0.0066. Bar indicates 100 μm. 

[Examination of the metaplastic milieu]

기존 연구에서 보면, SPEM에서 immune cells (특히, macrophages) infiltration없이 dysplasia가 발생하지 않습니다. 이번 연구에서도 CAG 에서 CD68 expressing macrophages의 infiltration이 현저하게 나타났으며 (Figure 3(c)) 특히, metaplastic glands 주위에 군집되어 나타났습니다. 반면에, AIG에서는 현저하게 적게 나타났으며 (Figure 3(d), 3 (e)), MPO-positive neutrophils에서도 적게 나타났습니다 (Figure 3(e)). Eosinophils의 경우 기존 연구에서는 AIG에서 높게 나타났으나, 이번 연구에서는 CAG에서 더 통계적으로 높게 나타났습니다 (위 table 1). 이러한 결과는, AIG 에서 proliferative metaplasia로 진행을 유발하는 현저한 macrophage infiltration이 매우 적다는 것을 의미합니다.

Figure 3. Characteristics of inflammatory infiltrates in metaplastic mucosa. Comparison of the infiltration of macrophages (CD68, green) and neutrophils (MPO, Myeloperoxidase, red) between chronic atrophic gastritis (CAG) ((a), (c)) and autoimmune gastritis (AIG) ((b), (d)) patients. Hematoxylin and eosin (H&E) staining of sections are shown from AIG (a) and CAG (b) patient stomach. In (c) and (d), spasmolytic polypeptide expressing metaplasia (SPEM) cells were stained with Griffonia simplicifolia II-lectin (white) and nuclei were stained with 40 ,6-diamidino-2-phenylindole (DAPI) (blue). Insets show higher magnification images as noted by white boxed outlines. Bar indicates 100 mm. (e) Quantitation of immunostaining demonstrated the presence of significantly fewer CD68-positive macrophages in AIG patients as compared to CAG patients (*p < 0.01).



Abstract 

Background: Autoimmune gastritis (AIG) and adenocarcinoma-associated chronic atrophic gastritis (CAG) are both associated with oxyntic atrophy, but AIG patients demonstrate an increased risk of carcinoid tumors rather than the elevated risk of adenocarcinoma observed with CAG. We therefore sought to compare the characteristics of the metaplastic mucosa in AIG and CAG patients. 

Methods: We examined markers for metaplasia (spasmolytic polypeptide expressing metaplasia (SPEM) and intestinal metaplasia) as well as proliferation (Ki67) and immune cell populations (neutrophils, macrophages, and eosinophils) in gastric sections from 16 female patients with autoimmune thyroiditis and AIG and 17 patients with CAG associated with gastric adenocarcinoma. 

Results: Both AIG and CAG patients demonstrated prominent SPEM and intestinal metaplasia. However, AIG patients displayed significantly lower numbers of infiltrating macrophages and significantly reduced mucosal cell proliferation as compared to CAG patients. 

Conclusions: These findings indicate that, while both AIG and CAG patients display prominent oxyntic atrophy and metaplasia, the AIG patients do not show proliferative metaplastic lineages that would predispose to adenocarcinoma.

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