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Regression of mucosa-associated lymphoid-tissue lymphoma after eradication of Helicobacter pylori
THE LANCET
that an autoimmune process also underlies the clinical condition in adults. Furthermore, because enterocyte antibodies were found in only two patients, Corazza and colleagues might know whether colitis was present in the patients with upper-intestinal disorder. Although the inflammatory process can be mild in the colon, most of the affected children with autoimmune enteropathy we have investigated presented with a generalised gut disorder of differing severity.3 Differences between the adult and the paediatric variant of autoimmune enteropathy might have already started to emerge. In fact, hyperplastic crypts and increased intraepithelial lymphocyte (IEL) counts were observed in the small-intestinal biopsies of the two reported adult cases, whereas in the paediatric samples hypoplastic crypts were found in the two original cases, and IEL counts were mainly decreased when a cohort of children was investigated.5 Finally, since enterocyte autoantibodies are routinely tested in human blood group O duodenum for diagnostic purposes, it would be of interest to know whether the same autoantibodies detected in the adult patients on monkey substrate also react with the human duodenum. Standard protocols should be undertaken and exchanges of serum samples among centres is needed to validate the detection of the enterocyte autoantibodies. *R Mirakian, P Collins, G F Bottazzo *Department of Immunology, St Bartholomews and Royal London School of Medicine and Dentistry, London E1 2AD, UK
1
2
3
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5
Corazza GR, Biagi F, Volta U, Andreani ML, De Franceschi L, Gasbarrini G. Autoantimmune enteropathy and villous atrophy in adults. Lancet 1997; 350: 106–09. Mirakian R, Richardson A, Milla PJ, et al. Protracted diarrhoea of infancy: evidence in support of an autoimmune variant. BMJ 1986; 293: 1132–36. Hill SM, Milla PJ, Bottazzo GF, Mirakian R. Autoimmune enteropathy and colitis: is there a generalised disorder? Gut 1991; 32: 36–42. Savage MO, Mirakian R, Wozniak ER, et al. Specific autoantibodies to gut epithelium in two infants with severe protracted diarrhoea. J Pediatr Gastroenterol Nutr 1985; 4: 187–95. Mirakian R, Hill S, Richardson A, Milla PJ, Walker-Smith JA, Bottazzo GF. HLA product expression and lymphocyte subpopulations in jejunum biopsies of children with idiopathic protracted diarrhoea and enterocyte autoantibodies. J Autoimmun 1988; 1: 263–77.
Authors’ reply SIR—We have reviewed the notes of our two adult patients with auto-
960
immune enteropathy. Colonoscopy had been done and multiple biopsies were taken. In both patients the large intestine was normal on endoscopy but the colonic biopsies showed an increase in intraepithelial lymphocytes (26% in patient 1; 49% in patient 2), some epithelial flattening, and absence of subepithelial collagen layer. Thus, according to Lazenby and colleagues 1 our patients also had lymphocytic colitis. We agree that these two cases lend further support to the hypothesis of a generalised gut disorder in patients who have autoimmune enteropathy . 2 Interestingly, adult patients with lymphocytic colitis, refractory coeliac disease, and associated autoimmune disease have been reported.3,4 Perhaps even those patients might have autoimmune enteropathy. As R Mirakian and colleagues point out, the study of the adult form of autoimmune enteropathy has just begun. More data will need to be collected before the adult variant can be compared with the paediatric one. Direct immunofluorescence of small-bowel biopsies and counting of intraepithalial / T-cells are among the first studies that should be carried out. As far as detection of enterocyte autoantibodies in human blood group O duodenum is concerned, the fact that they have been found even on rat intestine5 leads us to think that multiple species specificity for the enterocyte autoantibodies should be tested before any conclusion about optimum conditions. We agree with Mirakian and coworkers that this could be done by validation of enterocyte autoantibody detection through exchange of serum samples between centres. F Biagi, *G R Corazza Department of Internal Medicine, University of L’Aquila, 67100 L’Aquila, Italy
1
2
3
4
5
Lazenby AJ, Yardley JH, Giardiello FM, Jessurun J, Bayless TM. Lymphocytic (“microscopic”) colitis: a comparative histopathologic study with particular reference to collagenous colitis. Hum Pathol 1989; 20: 18–28. Hill SM, Milla PJ, Bottazzo GF, Mirakian R. Autoimmune enteropathy and colitis: is there a generalised autoimmune gut disorder? Gut 1991; 32: 36–42. Giardiello FM, Lazenby AJ, Bayless TM, et al. Lymphocytic (microscopic) colitis: clinicopathologic study of 18 patients and comparison to collagenous colitis. Dig Dis Sci 1989; 34: 1730–38. DuBois RN, Lazenby AJ, Yardley JH, et al. Lymphocytic enterocolitis in patients with “refractory sprue”. JAMA 1989; 262: 935–37. Walker-Smith JA, Unsworth DJ, Hutchins P, Phillips AD, Holborow EJ. Autoantibodies against gut epithelium in child with small-intestinal enteropathy. Lancet 1982; i: 566–67.
Regression of mucosa-associated lymphoid-tissue lymphoma after eradication of Helicobacter pylori SIR—Takayuki Matsumoto and colleagues (July 12, p 115)1 report a case of mucosa-associated lymphoid tissue (MALT) lymphoma of the rectum that regressed after treatment of Helicobacter pylori with omeprazole, amoxycillin, and clarithromycin. They note previous reports of regression of a salivery gland and small-intestinal MALT lymphomas in response to antihelicobacter therapy. Wotherspoon and co-workers2 originally showed that eradication of H pylori was followed by regression of gastric MALT lymphomas. In-vitro studies suggested that the proliferation of low-grade MALT lymphoma cells is driven by Tcells responding to specific H pylori derived antigens, and other investigators3 have shown that this is due to CD40-mediated signalling and Th2-type cytokines.3 Since H pylori colonises only gastrictype mucosa, how can treatment of this organism lead to regression of lymphomas at other sites? The case reported by Fischbach et al4 showed gastric metaplasia with H pylori infection in the duodenal biopsy and might, therefore, be expected to respond to therapy. It is possible that the other patients originally developed gastric MALT lymphomas that subsequently disseminated to non-gastric sites. At these sites, the lymphoma might receive the necessary drive either from H pylori antigens in the blood, or within the intestinal lumen, or from circulating T-cells that have been activated by H pylori antigens in the stomach. Dissemination of gastric MALT lymphoma to the intestine has been demonstrated,5 and is in keeping with the behaviour of lymphoid cells of the mucosal immune system. However, this explanation would imply that the primary gastric lymphoma in these patients was subclinical or had regressed before the patient was investigated. These reports do not, however, necessarily imply an association between H pylori and extra-gastric MALT lymphomas. The antibiotics used to eliminate H pylori are active against other organisms and conceivably might eliminate as yet unidentified agents responsible for the induction and growth of MALT lymphomas outside the stomach. D H Wright Southampton General Hospital, Southampton, Hampshire SO16 6YD, UK
1
Matsumoto T, Iida M, Shimizu M. Regression of mucosa-associated lymphoid-
Vol 350 • September 27, 1997
THE LANCET
2
3
4
5
tissue lymphoma of rectum after eradication of Helicobacter pylori. Lancet 1997; 350: 115–16. Wotherspoon AC, Doglioni C, Diss TC, et al. Regression of primary low-grade B-cell gastric lymphoma of mucosa-associated lymphoid tissue type after eradication of Helicobacter pylori. Lancet 1993; 342: 575–77. Greiner A, Knörr C, Qin Y, et al. Low-grade B-cell lymphomas of mucosa-associated lymphoid tissue (MALT-type) require CD40-mediated signalling and Th2-type cytokines for in vitro growth and differentiation. Am J Pathol 1997; 150: 1583–93. Fischbach W, Tacke W, Greiner A, Müller-Hermelink HK. Regression of immuno-proliferative small intestinal disease after eradication of Helicobacter pylori. Lancet 1997; 349: 31–32. Du MK, Xu CF, Diss TC, et al. Intestinal dissemination of gastric mucosa-associated lymphoid tissue lymphoma. Blood 1996; 88: 4445–51.
Authors’ reply SIR—Although regression of mucosaassociated lymphoid tissue (MALT) lymphoma outside the stomach after eradication of Helicobacter pylori suggests a possible association of the micro-organism with lymphoma genesis, we agree that more evidence is needed to confirm the hypothesis. Drobyski and Qazi1 found that a certain proportion of low-grade lymphomas shows spontaneous regression. Spontaneous regression has also been shown in follicular lymphoid hyperplasia of the intestine, a possible prototype of low-grade MALT lymphoma.2 Since the intestinal tract, especially the colorectum, is prone to various stimuli which can be altered by oral antibiotics,3 we cannot attribute solely to H pylori the regression of rectal MALT lymphoma in our patient. However, since previous viral infections or enhanced natural-killer activity, which are presumed to be critical in regression of lymphoma,1,4 were not evident in our case, it seems unreasonable to dissociate H pylori from the regression of rectal MALT lymphoma. The association of other microorganisms or agents seems unlikely, because no signs of recurrence were seen over 6 months even without any treatment. Without identification of other agents, H pylori can be proposed as a possible microorganism responisble for the pathogenesis of extragastric MALT lymphoma. *Takayuki Matsumoto, Mitsuo Iida, Michio Shimizu *Division of Gastroenterology and Department of Pathology, Kawasaki Medical School, Kurashiki City, Okayama 701-01, Japan
1
2
Drobyski WR, Qazi R. Spontaneous regression in non-Hodgkin’s lymphoma: clinical and pathogenetic considerations. Am J Hematol 1989; 31: 138–41. Ranchod M, Lewin KJ, Dorfman RF. Lymphoid hyperplasia of the gastrointestinal
Vol 350 • September 27, 1997
3
4
tract: a study of 26 cases and review of the literature. Am J Surg Pathol 1978; 2: 383–400. Midtvedt T, Carlstedt DB, Hoverstad T, Lingaas E, Norin E. Influence of peroral antibiotics upon the biotransformatory activity of the intestinal microflora in healthy subjects. Eur J Clin Invest 1986; 16: 11–17. Ono K, Kikuchi M, Funai N, Matsuzaki M, Shimamoto Y. Natural killing activity in patients with spontaneous regression of malignant lymphoma. J Clin Immunol 1996; 16: 334–39.
Use of anucleate donor oocyte cytoplasm in recipient eggs SIR—Jacques Cohen and colleagues (July 19, p 186)1 describe a successful birth after transfer of anucleate donor oocyte cytoplasm into recipient eggs. We welcome such reports, but would draw the investigators’ attention to several issues. They fail to mention whether donor mitochondria were excluded before the transfer and do not specify the amount of cytoplasm in proportion to total cytoplasm that was actually transferred. Indeed, they compare DNA fingerprinting profiles on amniocytes to establish genetic identity, but do not describe the degree of accuracy. This information is vital because very low levels of paternal mitochondrial DNA (mtDNA) inheritance have been detected in animals: paternal mtDNA has been detected at a frequency of 104 in interspecific crosses of mice.2 There are several important consequences related to this technique. The first is that should donor cytoplasm be transmitted as a result of this procedure, then the offspring would be the recipient of three genomes: the nuclear complements from the mother and father and some or all mitochondrial complement from the donor. If this occurred then several vital questions need to be answered. Male patients who have injections of intracytoplasmic sperm are recommended to undergo genetic screening. MtDNA point mutations and deletions are associated with several rare but life-threatening diseases. Importantly, these mutations are maternally inherited and indeed it is the ratio of wild type to deleted that would predict the pathogenicity of the disease. We thus suggest that the donor should, in this instance, be screened for possible carrier status, as is the case for potential donors of spermatozoa. Additionally, although mtDNA is not perceived to be the principal genome, it is vital to cell survival through the subunits it contributes to the process of oxidative phosphorylation. We have argued3 that in the ageing oocyte, in which mtDNA deletions are
high, a fault in the genetic filter, which would normally exclude deleted mtDNA, could either result in the transmission of deleted molecules or the possibility of paternal leakage or, in this case, donor mtDNA. The nucleus seems to be the regulator of mtDNA inheritance and transmission. Holt and colleagues 4 have shown, in two separate cell lines, that mtDNA is regulated by the cell’s nuclear background, with one line eradicating the mutation in favour of wild type and the other allowing the mutation to be further continued during passaging. The question that needs to be answered is whether the oocyte nuclear background remains the same in all cases or when it is partly disrupted does its nuclear selection have less influence. In the procedure used by Cohen, it is highly likely that because of oocyte factors that influence the fertility of the couple, damage to the oocyte could affect cytoplasmic selection. We recommend that if cytoplasmic donation is to become clinical practice, as suggested by Kahn et al,5 for women who harbour mtDNA deletions, then more experimental work needs to be undertaken. *Justin C St John, Christopher L R Barratt Department of Medicine, University of Birmingham, and Birmingham Women’s Hospital, Birmingham B15 2TG, UK
1
2
3
4
5
Cohen J, Scott R, Schimmel T, Levron J, Willadsen S. Birth of infant after transfer of anucleate donor oocyte cytoplasm into recipient eggs. Lancet 1997; 350: 186–87. Gyllesten U, Wharton D, Josefsson A, Wilson AC. Paternal inheritance of mitochondrial DNA in mice. Nature 1991; 353: 255–57. St John JC, Cooke ID, Barratt CLR. Mitochondrial mutations and male infertility. Nature Med 1997; 3: 124–25. Holt IJ, Dunbar DR, Jacobs HT. Behaviour of a population of partially duplicated mitochondrial DNA molecules in cell culture: segregation, maintenance and recombination dependent upon nuclear background. Human Mol Genet 1997; 6: 1251–60. Kahn A. Clone mammals: clone man? Nature 1996; 386: 119.
SIR—Justin St John and Christopher Barratt raise interesting issues about the transfer of mtDNA into developmentally-jeopardised oocytes. Cell-cycle mechanisms as well as injected mRNA seem to have a pivotal role in accomplishing a successful turnaround in ooplasmic and nuclear competence. The immediate role of a foreign pool of mitochondira is hypothetical. The cytoplasmic transfer procedure used in our protocol does not exclude any particular organelle apart from the nucleus. About 5% of ooplasm was transferred in our patient; and it is fairly easy to replace all or large proportions. The small amount
961
that an autoimmune process also underlies the clinical condition in adults. Furthermore, because enterocyte antibodies were found in only two patients, Corazza and colleagues might know whether colitis was present in the patients with upper-intestinal disorder. Although the inflammatory process can be mild in the colon, most of the affected children with autoimmune enteropathy we have investigated presented with a generalised gut disorder of differing severity.3 Differences between the adult and the paediatric variant of autoimmune enteropathy might have already started to emerge. In fact, hyperplastic crypts and increased intraepithelial lymphocyte (IEL) counts were observed in the small-intestinal biopsies of the two reported adult cases, whereas in the paediatric samples hypoplastic crypts were found in the two original cases, and IEL counts were mainly decreased when a cohort of children was investigated.5 Finally, since enterocyte autoantibodies are routinely tested in human blood group O duodenum for diagnostic purposes, it would be of interest to know whether the same autoantibodies detected in the adult patients on monkey substrate also react with the human duodenum. Standard protocols should be undertaken and exchanges of serum samples among centres is needed to validate the detection of the enterocyte autoantibodies. *R Mirakian, P Collins, G F Bottazzo *Department of Immunology, St Bartholomews and Royal London School of Medicine and Dentistry, London E1 2AD, UK
1
2
3
4
5
Corazza GR, Biagi F, Volta U, Andreani ML, De Franceschi L, Gasbarrini G. Autoantimmune enteropathy and villous atrophy in adults. Lancet 1997; 350: 106–09. Mirakian R, Richardson A, Milla PJ, et al. Protracted diarrhoea of infancy: evidence in support of an autoimmune variant. BMJ 1986; 293: 1132–36. Hill SM, Milla PJ, Bottazzo GF, Mirakian R. Autoimmune enteropathy and colitis: is there a generalised disorder? Gut 1991; 32: 36–42. Savage MO, Mirakian R, Wozniak ER, et al. Specific autoantibodies to gut epithelium in two infants with severe protracted diarrhoea. J Pediatr Gastroenterol Nutr 1985; 4: 187–95. Mirakian R, Hill S, Richardson A, Milla PJ, Walker-Smith JA, Bottazzo GF. HLA product expression and lymphocyte subpopulations in jejunum biopsies of children with idiopathic protracted diarrhoea and enterocyte autoantibodies. J Autoimmun 1988; 1: 263–77.
Authors’ reply SIR—We have reviewed the notes of our two adult patients with auto-
960
immune enteropathy. Colonoscopy had been done and multiple biopsies were taken. In both patients the large intestine was normal on endoscopy but the colonic biopsies showed an increase in intraepithelial lymphocytes (26% in patient 1; 49% in patient 2), some epithelial flattening, and absence of subepithelial collagen layer. Thus, according to Lazenby and colleagues 1 our patients also had lymphocytic colitis. We agree that these two cases lend further support to the hypothesis of a generalised gut disorder in patients who have autoimmune enteropathy . 2 Interestingly, adult patients with lymphocytic colitis, refractory coeliac disease, and associated autoimmune disease have been reported.3,4 Perhaps even those patients might have autoimmune enteropathy. As R Mirakian and colleagues point out, the study of the adult form of autoimmune enteropathy has just begun. More data will need to be collected before the adult variant can be compared with the paediatric one. Direct immunofluorescence of small-bowel biopsies and counting of intraepithalial / T-cells are among the first studies that should be carried out. As far as detection of enterocyte autoantibodies in human blood group O duodenum is concerned, the fact that they have been found even on rat intestine5 leads us to think that multiple species specificity for the enterocyte autoantibodies should be tested before any conclusion about optimum conditions. We agree with Mirakian and coworkers that this could be done by validation of enterocyte autoantibody detection through exchange of serum samples between centres. F Biagi, *G R Corazza Department of Internal Medicine, University of L’Aquila, 67100 L’Aquila, Italy
1
2
3
4
5
Lazenby AJ, Yardley JH, Giardiello FM, Jessurun J, Bayless TM. Lymphocytic (“microscopic”) colitis: a comparative histopathologic study with particular reference to collagenous colitis. Hum Pathol 1989; 20: 18–28. Hill SM, Milla PJ, Bottazzo GF, Mirakian R. Autoimmune enteropathy and colitis: is there a generalised autoimmune gut disorder? Gut 1991; 32: 36–42. Giardiello FM, Lazenby AJ, Bayless TM, et al. Lymphocytic (microscopic) colitis: clinicopathologic study of 18 patients and comparison to collagenous colitis. Dig Dis Sci 1989; 34: 1730–38. DuBois RN, Lazenby AJ, Yardley JH, et al. Lymphocytic enterocolitis in patients with “refractory sprue”. JAMA 1989; 262: 935–37. Walker-Smith JA, Unsworth DJ, Hutchins P, Phillips AD, Holborow EJ. Autoantibodies against gut epithelium in child with small-intestinal enteropathy. Lancet 1982; i: 566–67.
Regression of mucosa-associated lymphoid-tissue lymphoma after eradication of Helicobacter pylori SIR—Takayuki Matsumoto and colleagues (July 12, p 115)1 report a case of mucosa-associated lymphoid tissue (MALT) lymphoma of the rectum that regressed after treatment of Helicobacter pylori with omeprazole, amoxycillin, and clarithromycin. They note previous reports of regression of a salivery gland and small-intestinal MALT lymphomas in response to antihelicobacter therapy. Wotherspoon and co-workers2 originally showed that eradication of H pylori was followed by regression of gastric MALT lymphomas. In-vitro studies suggested that the proliferation of low-grade MALT lymphoma cells is driven by Tcells responding to specific H pylori derived antigens, and other investigators3 have shown that this is due to CD40-mediated signalling and Th2-type cytokines.3 Since H pylori colonises only gastrictype mucosa, how can treatment of this organism lead to regression of lymphomas at other sites? The case reported by Fischbach et al4 showed gastric metaplasia with H pylori infection in the duodenal biopsy and might, therefore, be expected to respond to therapy. It is possible that the other patients originally developed gastric MALT lymphomas that subsequently disseminated to non-gastric sites. At these sites, the lymphoma might receive the necessary drive either from H pylori antigens in the blood, or within the intestinal lumen, or from circulating T-cells that have been activated by H pylori antigens in the stomach. Dissemination of gastric MALT lymphoma to the intestine has been demonstrated,5 and is in keeping with the behaviour of lymphoid cells of the mucosal immune system. However, this explanation would imply that the primary gastric lymphoma in these patients was subclinical or had regressed before the patient was investigated. These reports do not, however, necessarily imply an association between H pylori and extra-gastric MALT lymphomas. The antibiotics used to eliminate H pylori are active against other organisms and conceivably might eliminate as yet unidentified agents responsible for the induction and growth of MALT lymphomas outside the stomach. D H Wright Southampton General Hospital, Southampton, Hampshire SO16 6YD, UK
1
Matsumoto T, Iida M, Shimizu M. Regression of mucosa-associated lymphoid-
Vol 350 • September 27, 1997
THE LANCET
2
3
4
5
tissue lymphoma of rectum after eradication of Helicobacter pylori. Lancet 1997; 350: 115–16. Wotherspoon AC, Doglioni C, Diss TC, et al. Regression of primary low-grade B-cell gastric lymphoma of mucosa-associated lymphoid tissue type after eradication of Helicobacter pylori. Lancet 1993; 342: 575–77. Greiner A, Knörr C, Qin Y, et al. Low-grade B-cell lymphomas of mucosa-associated lymphoid tissue (MALT-type) require CD40-mediated signalling and Th2-type cytokines for in vitro growth and differentiation. Am J Pathol 1997; 150: 1583–93. Fischbach W, Tacke W, Greiner A, Müller-Hermelink HK. Regression of immuno-proliferative small intestinal disease after eradication of Helicobacter pylori. Lancet 1997; 349: 31–32. Du MK, Xu CF, Diss TC, et al. Intestinal dissemination of gastric mucosa-associated lymphoid tissue lymphoma. Blood 1996; 88: 4445–51.
Authors’ reply SIR—Although regression of mucosaassociated lymphoid tissue (MALT) lymphoma outside the stomach after eradication of Helicobacter pylori suggests a possible association of the micro-organism with lymphoma genesis, we agree that more evidence is needed to confirm the hypothesis. Drobyski and Qazi1 found that a certain proportion of low-grade lymphomas shows spontaneous regression. Spontaneous regression has also been shown in follicular lymphoid hyperplasia of the intestine, a possible prototype of low-grade MALT lymphoma.2 Since the intestinal tract, especially the colorectum, is prone to various stimuli which can be altered by oral antibiotics,3 we cannot attribute solely to H pylori the regression of rectal MALT lymphoma in our patient. However, since previous viral infections or enhanced natural-killer activity, which are presumed to be critical in regression of lymphoma,1,4 were not evident in our case, it seems unreasonable to dissociate H pylori from the regression of rectal MALT lymphoma. The association of other microorganisms or agents seems unlikely, because no signs of recurrence were seen over 6 months even without any treatment. Without identification of other agents, H pylori can be proposed as a possible microorganism responisble for the pathogenesis of extragastric MALT lymphoma. *Takayuki Matsumoto, Mitsuo Iida, Michio Shimizu *Division of Gastroenterology and Department of Pathology, Kawasaki Medical School, Kurashiki City, Okayama 701-01, Japan
1
2
Drobyski WR, Qazi R. Spontaneous regression in non-Hodgkin’s lymphoma: clinical and pathogenetic considerations. Am J Hematol 1989; 31: 138–41. Ranchod M, Lewin KJ, Dorfman RF. Lymphoid hyperplasia of the gastrointestinal
Vol 350 • September 27, 1997
3
4
tract: a study of 26 cases and review of the literature. Am J Surg Pathol 1978; 2: 383–400. Midtvedt T, Carlstedt DB, Hoverstad T, Lingaas E, Norin E. Influence of peroral antibiotics upon the biotransformatory activity of the intestinal microflora in healthy subjects. Eur J Clin Invest 1986; 16: 11–17. Ono K, Kikuchi M, Funai N, Matsuzaki M, Shimamoto Y. Natural killing activity in patients with spontaneous regression of malignant lymphoma. J Clin Immunol 1996; 16: 334–39.
Use of anucleate donor oocyte cytoplasm in recipient eggs SIR—Jacques Cohen and colleagues (July 19, p 186)1 describe a successful birth after transfer of anucleate donor oocyte cytoplasm into recipient eggs. We welcome such reports, but would draw the investigators’ attention to several issues. They fail to mention whether donor mitochondria were excluded before the transfer and do not specify the amount of cytoplasm in proportion to total cytoplasm that was actually transferred. Indeed, they compare DNA fingerprinting profiles on amniocytes to establish genetic identity, but do not describe the degree of accuracy. This information is vital because very low levels of paternal mitochondrial DNA (mtDNA) inheritance have been detected in animals: paternal mtDNA has been detected at a frequency of 104 in interspecific crosses of mice.2 There are several important consequences related to this technique. The first is that should donor cytoplasm be transmitted as a result of this procedure, then the offspring would be the recipient of three genomes: the nuclear complements from the mother and father and some or all mitochondrial complement from the donor. If this occurred then several vital questions need to be answered. Male patients who have injections of intracytoplasmic sperm are recommended to undergo genetic screening. MtDNA point mutations and deletions are associated with several rare but life-threatening diseases. Importantly, these mutations are maternally inherited and indeed it is the ratio of wild type to deleted that would predict the pathogenicity of the disease. We thus suggest that the donor should, in this instance, be screened for possible carrier status, as is the case for potential donors of spermatozoa. Additionally, although mtDNA is not perceived to be the principal genome, it is vital to cell survival through the subunits it contributes to the process of oxidative phosphorylation. We have argued3 that in the ageing oocyte, in which mtDNA deletions are
high, a fault in the genetic filter, which would normally exclude deleted mtDNA, could either result in the transmission of deleted molecules or the possibility of paternal leakage or, in this case, donor mtDNA. The nucleus seems to be the regulator of mtDNA inheritance and transmission. Holt and colleagues 4 have shown, in two separate cell lines, that mtDNA is regulated by the cell’s nuclear background, with one line eradicating the mutation in favour of wild type and the other allowing the mutation to be further continued during passaging. The question that needs to be answered is whether the oocyte nuclear background remains the same in all cases or when it is partly disrupted does its nuclear selection have less influence. In the procedure used by Cohen, it is highly likely that because of oocyte factors that influence the fertility of the couple, damage to the oocyte could affect cytoplasmic selection. We recommend that if cytoplasmic donation is to become clinical practice, as suggested by Kahn et al,5 for women who harbour mtDNA deletions, then more experimental work needs to be undertaken. *Justin C St John, Christopher L R Barratt Department of Medicine, University of Birmingham, and Birmingham Women’s Hospital, Birmingham B15 2TG, UK
1
2
3
4
5
Cohen J, Scott R, Schimmel T, Levron J, Willadsen S. Birth of infant after transfer of anucleate donor oocyte cytoplasm into recipient eggs. Lancet 1997; 350: 186–87. Gyllesten U, Wharton D, Josefsson A, Wilson AC. Paternal inheritance of mitochondrial DNA in mice. Nature 1991; 353: 255–57. St John JC, Cooke ID, Barratt CLR. Mitochondrial mutations and male infertility. Nature Med 1997; 3: 124–25. Holt IJ, Dunbar DR, Jacobs HT. Behaviour of a population of partially duplicated mitochondrial DNA molecules in cell culture: segregation, maintenance and recombination dependent upon nuclear background. Human Mol Genet 1997; 6: 1251–60. Kahn A. Clone mammals: clone man? Nature 1996; 386: 119.
SIR—Justin St John and Christopher Barratt raise interesting issues about the transfer of mtDNA into developmentally-jeopardised oocytes. Cell-cycle mechanisms as well as injected mRNA seem to have a pivotal role in accomplishing a successful turnaround in ooplasmic and nuclear competence. The immediate role of a foreign pool of mitochondira is hypothetical. The cytoplasmic transfer procedure used in our protocol does not exclude any particular organelle apart from the nucleus. About 5% of ooplasm was transferred in our patient; and it is fairly easy to replace all or large proportions. The small amount
961