TPC NEWS Vol.4, No.2, Fall 1985, (Whole Number 8)

( English Summary )

 

Page-3 Essay: Coexistence with aged monkeys

     As of June of this year, the oldest cynomolgus monkeys, who are being kept at TPC and whose exact birth dates are known, are 22 years and four months old with male, 20 years and six months with female.  Regarding wild-originated breeders, the oldest female and male are estimated to be 22 to 25 years of age.  It is generally accepted that the life span of the macaque would be 25 to 30 years.  Thus our old monkeys may be worthy of being celebrated for their long lives.

     At TPC, we intend not only to hold a very large number of young monkeys but also to keep as many 'aged monkeys as possible, because we think that old monkeys are able to b" a suitable animal model for studies aging and senility in man.

   In general, the orthodox method of animal experiment in studies of aging and senility are to observe and measure natural changes with increasing ages in morphology, functions, behaviors and some reactions to a certain treatment. To adopt this method of study is rather easy, when we deal with laboratory animals of comparatively short life span, such as mice. Usually, however, we can not apply this method to animals of long life like primates. It is almost impossible to perform follow-up studies on such animals throughout their lives, in other words, to adopt a longitudinal method of study, judging from ordinary duration of research activities of a scientist. So it is impossible for us not to take a transverse way of study in which we employ animals of different ages at a certain moment.

     Naturally, this method of study would require maintaining and conditioning many monkeys of different age classes. To satisfy this prerequisite is rather difficult for ordinary laboratories or facilities of experimental animals in universities and research institutes.  I think, fortunately, TPC has this prerequisite at the present time.  But to keep this condition from now on is not easy because it needs special care, area, budget and patience.

    By the way, I, being in..1%1 late. fifties, have begun to feel my own "age".  Although I keenly feel that more much time is necessary for me as a scientist to develop my work, 'I will have to retire from TPC within five years.  Therefore, I wish that TPC could keep those aged monkeys safely as long as possible and I make their biological characteristics as clear as possible.  These works will remain to my successors.

     If the young staffs of this center who are and will be working with aged monkeys would have thought about their own way of life through cares or works on aged monkeys, in addition to satisfying their will of scientific research, then it would be my great pleasure.

 

Page-4 Breeding Topics: The third primate species kept at TPC---Common squirrel monkey

    The common squirrel monkey is a species of the New World monkey whereas both the cynomolgus and the green monkeys are 01d World monkeys. The latter two species live on the ground but the squirrel monkey lives usually in trees. Adult female squirrel monkeys weigh 600-800g and males about 1kg.  Birth weight is around 100g. They are usually have one baby at a time.

The nursing behavior of this species is unique. Mother monkeys carry their babies on the back.  When they suckle they draw their babies to their breasts.  They scarcely hold babies in their arms even when they give milk.

    Unlike the cynomolgus and green monkeys , the squirrel monkey has seasonality of breeding.  They mate from late fall to winter it the northern hemisphere and have babies from spring to early summer.  Many studies have been done on the breeding seasonality by using squirrel monkeys.  But the exact mechanism to provoke this seasonality is still unknown .

    At present, the squirrel monkeys kept at TPC are 9 adult males and 39 females.  They were imported during from Feb. '80 to Nov. ' 82.  Until today 31 male and 16 female offsprings were born in this center.  Since we introduced squirrel monkeys to our colony, we have maintained the species on the basis of the breeding systems established for the cynomolgus monkey. The breeding performance are shown in the table below.  The items given are, from the top column to the bottom, (1) the period during which we found the pregnancy, (2) the period of delivery, (3) the number of delivery to the number of female breeders.  The number of delivery has surely increased.  However, further investigations are needed to determine whether or not the seasonality has become less marked for these five years.

    Colony bred squirrel monkeys born in 1981 has been fertile. We  have already gotten 27 infants up to August 7 of this year.

 

Page-5 A Survey on the Incidence of Antibody to Simian T-Cell Leukemia Virus in Cynomolgus and Green Monkeys

    Retrovirus having close antigenic similarity to Adult-T-Cell Leukemia Virus (ATLV or HTLV-Type 1) has been found in various 01d World monkey species .  It has been proposed to call the virus Simian T-Cell Leukemia Virus (STLV) .

   Surveys on the incidence of an antibody to STLV by the indirect immuno-fluorescence method were conducted with wild originated monkeys.

[ Materials and methods ]

     Sera used were collected from 651 cynomolgus monkeys of wild origin which were imported from South-east Asia in 1968 to 1982 and thereafter kept in our center, 150 newly imported cynomolgus monkeys from the Philippines, 45 African green monkeys of feral origin imported in 1979 to 1981 and one to four years old 69 offsprings born from these imported wild green monkeys .

     The antigen used was MT-1 cell established by Miyoshi et, al.  MT-1 cells spread on slide glasses were dried and treated with acetone for 10 mmutes at room temperature .

   FITC labeled anti-monkey IgG goat serum diluted 170 times was used as the second serum.

[ Result ]

Wild Originated Cynomolgus Monkeys:  Of 651 animals maintained for a long period, 76(11.7%) were positive.  According to their countries of origin, 52(23.9%) of 218 Indonesian monkeys, 22(9.4%) of 233 Malaysian monkeys and 1(0.5%) of 198 Philippine monkeys were positive. A Cambodian monkey, though there was only one in our colony, was positive .  But, one animal from Thailand was negative .

Newly Imported Cynomolgus Monkeys :  Thirteen (8.7%) of 150 animals were positive.

African Green Monkeys of Wild Origin: Thirty-two(70%) of 46 animals were positive.  They had already been positive at the time of arrival at TPC.

Colony-bred African Green Monkeys: All 69 animals were negative. .

[Conclusion]

    As for the wild originated cynomolgus monkeys kept for a long time, the positive rate differed with the country of origin, But further studies are needed to determine whether or not these differences show the real state of STLV-contamination in those countries. Definite reasons for the large difference in the antibody incidence between newly imported Philippine monkeys and long-maintained Philippine monkeys is unknown at the present moment.

    High positive rate of the wild originated African green monkeys indicates that when we handle African green monkeys we should be more careful from the standpoint of biohazard control, It is still unclear why all the colony-bred and -reared green monkeys were negative, It may be considered that these monkeys were still too young or there is no virus transmission from mother monkeys.  Periodical examinations are needed to clear these problems.

 

Page-6 Introduction of Facilities - Facility for incineration

     At TPC sewage water from animal rooms amounts to 2000 mg a day, and drainage from research laboratories amounts to 20 m a day. We have a sewage treatment facility (See, TPC NEWS Vol., 1 No. 1 ) and a facility for incineration system to treat the leftovers and excrement separated from the sewage water by automatic scriveners.

    In the incineration building, there are a main incinerator and a sub-incinerator. The main incinerator has two furnaces. One of the furnaces is for combustion and the other for re-combustion. The sub-incinerator is used to dispose of ordinary rubbish. The combustion furnace of the main incinerator is 1.9m wide, 2.9m deep and 1.9 high.  The chamber of this furnace is equipped with four jet burners to get the best combustion efficiency.  The inner wall of the furnace on which temperature comes to 700Ž is made of firebrick and the outer side is covered with heat-proof bricks.  On the ceiling, a way-in is fixed with a dumper. The re-combustion furnace having two jet burners is for burning up odors and gasses resulting from imperfect combustion in the furnace I mentioned above.

  Solid filth is carried by a skip conveyor to the way-in on the upper surface of the incinerator and falls into the furnace.  Carcasses of autopsied animals are also incinerated in this furnace.  They are thrown into the furnace through the other door fixed on the side wall. The sub-incinerator has no burner, however, once the rubbish is kindled it is  completely burned up owing to a chimney eleven meters high.

   These incinerators have worked well without any trouble and accident since they were installed seven years ago.  This is due to earnest efforts of maintenance,

    Moreover, we have adopted a room of 28 m3 to keep solid waste or carcasses of animals at 5Ž for the occasion sit which the incinerator does not work.

 

Page-7 Homeland of the Cynomolgus Monkey - My field study of the cynomolgus monkey in Indonesia

     The time when I began to deal with cynomolgus monkeys in my research work was coincident with the time when TPC was established, through the process of studies on phylogeny of the Japanese monkey by the method of population genetics.  I became aware of the usefulness of cynomolgus monkeys, especially the ones living in Indonesia as important research materials.

     For these two decades, knowledge concerning genetic variation of natural population has been rapidly accumulated owing to the advanced technique of electrophoresis.  Many series of studies are being carried out with various animal species in order to elucidate a number of problems such as maintenance mechanism of intra-species variation, population structure, specific divergence and phylogenetic relations.

    The results obtained from the studies using Japanese monkeys have revealed that (1) a troop as a social unit of the Japanese macaque is open to other troops in respect of breeding, (2) the genetic variability within a troop is low and (3)the genetic differentiation between troops is limited.  Comparative studies using other primate species that are close relatives of the Japanese macaque and living in boundary islands in the distribution area of macaques were needed to determine whether or not those findings with the Japanese monkey were recognized also in other primate species.  That is the very reason why I chose the cynomolgus monkey in Indonesia as my research material.

    My field work needed capturing monkeys so that I could collect blood samples from troops of wild populations.  I began my research plan in seeking cynomolgus monkeys which were habituated to humans in Sumatra, Java and Bali Island.  Then I found that these three islands had their own characteristics in the relationship between monkeys and men.

    For example, in Sumatra Island, even though vast tropical forest is still existing, cynomolgus monkeys are now exposed to serious environmental conditions such as destruction of the forest by road constructions and capture for exportation. The living condition in this island is so severe that people seemed not to have any surplus to feed monkeys.  Therefore, it was so difficult to find cynomolgus troops composed with habituated monkeys.  However, harvesting bananas by using trained pig-tail monkeys was a memorable custom in Minancabau district in the state of West Sumatra. I asked people whether cynos were able to be trained for such a custom.  They replied that pig-tails were wiser than cynos, This reply hurt my feeling a little because I had been attached to cynos.

    Java Island is one of the most overpopulated zones in the world. Differing from the sight of Sumatra, I could find inhabited localities in any mountainous place of Java far from the highway.

   In Sunda Islands, east of Sunda Strait, the cynomolgus monkey is only one species of the genus Macaca.  Although Java is an overpopulated area, I had many chance to meet cynos there.  Even in Jakarta, the capital of six million population, I saw cynomolgus monkeys living vigorously in the mangrove forest. The cynos which live under symbiotic conditions with man reminded me of a scientist's comment that the environmental adaptability of the genus Macaca can be expressed by the words, "weed-like nature'.

    On the other hand, Bali is an island of Hindus and rice fields.  Temples called Pula are in villages of Bali.  In the woods surrounding those temples I often saw cynomolgus monkeys.  Villagers' feelings seemed not to be hostile to monkeys in spite of damages caused by monkeys in rice fields.  It may be because that people have respect for Hanuman, a monkey god.  There even are some Hindu temples that are famous for monkeys.  The situation in Bali which monkeys and men live in peace with religious background was utterly different from that in Sumatra and Java Island.

    Then I visited Lombok and Sumbawa islands, but I could not have any chance to see "habituated" monkeys.  So I decided to carefully observe feral monkeys and to capture them, Although both Lombok and Sumbawa islands are dry districts, monkeys have adapted themselves to such bad natural environment. 

    As a result of surveys in these five islands, I could collect blood samples from 456 animals of 29 troops from 10 habitats.

 

    Electrophoretic analyses of 29 kinds of proteins which were considered to be controlled by 33 gene-loci demonstrated the presence of polymorphism with regard to 19 gene-loci.  Frequency distribution of certain genetic markers in several population categories showed that the troops of the cynomolgus monkey in Indonesia were open units in respect to breeding like the troops of the Japanese monkey.  Genetic exchange due to traveling of adult male monkeys among troops could be considered.  This supposition was also supported by the field observation in Sumatra and Borneo.

   On the other hand, quantitative analyses of genetic variability in the troops revealed their genetic similarity and dissimilarity to the Japanese monkey.  As shown in Table 1, the genetic differentiation of the troops of cynomolgus monkeys was three times higher than that of Japanese monkeyfs.  These results mean that the genetic structure of members in a cynomolgus troop is not uniform. Furthermore, the definitive point in differentiating them from the Japanese monkey is that a remarkable difference in genetic structure was observed among troops in different islands.

    Table II shows the structure of genetic divergence among the cynomolgus monkey populations.  Here, I supposed such a hierarchial population structure that the whole population is just a mass of insular populations and an insular population is just as a mass of troops.  The relative genetic divergence among insular populations was calculated based on the estimated genetic frequency of troops.  These results showed that the relative divergence of the cynomolgus monkey population was 41.3%, whereas that of the Japanese monkey population in Honshu, Shikoku and Kyushu was much lower, that is only 2.7%.  In short, my field studies in Indonesia have revealed that the cynomolgus monkey has genetic characteristics, that is the difference in genetic structure is large among different local populations in different islands.

    When we establish nonhuman primates as laboratory animals, the genetic differentiation of great magnitude is a very useful factor for using in various experiments.  Moreover, many genetic markers kept in nonhuman primates are expected to be a strong weapon for genetic analyses.

    On the other hand, when we notice the large genetic differentiation among local populations, breeding nonhuman primates with each group divided by their country of origin is of great significance in maintaining genetic variabilities and in doing phylogenetic studies.  Now, I am very interested in how "weed-like nature' of the cynomolgus monkey, with which I was strongly impressed in my field work, will be developed in the process of laboratory conditioning.

 

Page-9 A Comment on My Research - Experiences of biochemical examinations on the blood of cynomolgus monkeys

Foreword:

     In our breeding and rearing facilities, biochemical blood-examinations are carried out to grasp general health conditions of animals.  Now I am in charge biochemical serum examinations using an auto-analyzer.  As everybody knows, an auto-biochemical-analyzer is useful to obtain simultaneously and speedily many data, of many items using a minute quantity of sample.  But an auto-analyzer is only machine which calculates the data resulting from the reaction of reagents to test-materials.  Therefore, when we judge the health conditions of animals from the data, we have to know beforehand the normal changes with increasing age, differences between males and females, and so on.  In this paper, I will describe my experiences associated with blood-examinations of cynomolgus monkeys using an auto-analyzer,

1. The influence of freezing and thawing.

    At TPC, periodical health checkups are carried out on every Thursday morning. The blood taken is kept all night at 4Ž.Sera are separated the next morning and are analyzed in the afternoon of the same day. Routine analysis using an auto analyzer is performed once a week.  As for the animals suspected to be suffering from some diseases, their sera are examined also by some method other than the auto-analyzer. When we can not immediately examine the sera they are frozen and preserved until the next routine analysis.  Moreover, re-examination of preserved 't sera is sometimes needed.

    So I tried to examine the influences of freezing and thawing and of the preservation period.

    Sera from several male cynomolgus monkeys were mixed and then divided into 1 m1 aliquot, The items and the unit of measurement are shown in Table 1.  The results about GPT are given in Fig. 1.  Symbol o (solid line) shows the data taken after freeze-preservation at -804: and symbol + (interrupted line) at -20Ž.   A gradual decrease in GPT activity was observed during one-year freeze-preservation at -80Ž: and a very rapid decrease was observed at -20Ž.  GOT had little change at -80Ž: while it decreased during the long preservation .period at -20Ž ALP decreased gradua;1y after eight months regardless of the preservation temperature. TP, ALB, BUN, GLU and T-CHO showed no remarkable change throughout the preservation period of one year.  The values of F-CHO and TG tended to rise with the increase of the preservation period.  When we analyze frozen-preserved sera we have to be careful in the judgment of the data obtained with regard to some items.

 

2. The influence of concentration before measurement.

    When I was inexperienced in operating the auto-analyzer I often could not finish analyzing materials within a day after I got the materials.  Under such circumstances, I had to measure them again the next day.  And I found that the data obtained on the next day seemed to be a little higher than those obtained on the day before.  I thought that the cause to increase the value would be the concentration of the materials during the time before measurement.  I examined the influence of the concentration on the values when materials were left for a long time at room temperature.

   Sera were taken from ten animals.  Three aliquots of 300, alwere prepared with each serum.  One of them was analyzed on the day the blood was taken.  The rest aliquot samples were left for six hours at room temperature, and then kept at 4Ž.  On the next day, one of the rest aliquots was measured.  Another one was again left at room temperature and kept overnight at 4Ž: and then analyzed.  The results are given in Table 2.

    Of course, the concentration of test-materials can be influenced by the temperature and humidity of a room where samples are left.  Therefore, the data may be affected by the difference in type of the analyzer used as well as by season.  Now, that I have become accustomed to the operation of auto-analyzer, I make it a point to set a half or one-third of all of the materials in the tray and not to place them at a time when I measure a lot of samples.

3. The physiological adjustment of the wild originated animal to the artificial environment.

     One of the problems in wild originated animals is the physiological change resulting from their transference from a natural environment to an artificial one.  At TPC, the quarantine period of at least three months is set for them.  So I examined the changes of various physiological parameters in newly-imported cynomolgus monkeys during the time of arrival at TPC to the 3rd and 6th month after arrival.  Two typical changes are given in Figs. 2 (BUN) and 3 (ALB) .

     Generally, both male and female BUN were low on arrival, In some exceptional cases they-were high.  After three months, the animals showed neither an extremely high level nor an extremely low level Six months later the values rose slightly as a whole.  These changes suggest that the animals generally be-came stable in their physiologic state.   As for ALB, the changes of the values showed the same pattern as those of BUM.  It can be said that the newly imported monkeys including some with ill health conditions recovered three months later and adjusted themselves to the artificial environment after six months.

   As for serum Ca, I had an interesting experience related to its concentration.  Fig.3 shows the frequency distributions of serum Ca concentrations on arrival in four different groups of newly imported cynomolgus monkeys. These four groups were imported almost at the same time. Lot 10-3 differs from others in regard to the country of origin  and shows lower values.  The average estimated age of animals in Lot 10-3, judging from dentition, was younger than that in other lots and Lot 10-3 was lower in weight, too.  Therefore I examined the relations between serum Ca concentration and age and/or weight of animals, but I could not find any significant relation.  At the end of the quarantine period, however as I presented in Fig. 5, all animals including those of Lot 10-3 showed the values around 9.8 mg/d1. Any way, it is highly probable that the serum Ca level of imported wild monkeys can be improved by the feeding conditions of TPC during the quarantine period.

 

4. The relation between age and determined values in colony bred monkeys.

    It is important to know the relation between age and determined values.  I made transverse studies on the relation using 555 female and 531 male cynomolgus monkeys bred-and-reared in our colonies.

    The item that exhibited the most remarkable change was ALP(Fig. 6).  ALP of infant and juvenile monkeys showed both high activities and wide individual variations.  The activities, however, markedly decreased after the age of 4 to 5 years.  At the age of 6, the activity dropped to one- sixth of the activity at the age of one or two.  The activity declined a little earlier in females than in males.  As Figs. 6 and 7 show, the time when ALP became stable at low value and the time when the weight increase reached the plateau are almost coincident with each other.  These results suggest that the cynomolgus monkey is fully matured by the age of six. 

    By the way, it has been revealed by the examination of sex-steroid hormonesf level in blood that the sexual maturation of the cynomolgus monkeys is attained at the age of about 4.  Also in this respect, females mature earlier than males, and the time of maturation corresponds with the time of rapid decrease of ALP.

 

Afterword:

    The knowledge I described above resulted from the examinations performed with cynomolgus monkeys.  At TPC, African green monkeys and squirrel monkeys are kept besides cynomolgus monkeys.  These two additional species are now small in number, but we will be able to carry out the same examinations using them in a few years.

    I have performed several basic examinations to obtain reliable data by which we can grasp health conditions of animals.  From now on I constantly have to make every effort to re-examine the conditions for measurement as well as examination items themselves.

 

Page-13 A Special Article - Studies on basement membranes and laminin components - Usefulness of the nonhuman primate.

    During the past decade the study of the chemistry and biology of basement membranes has been influenced by the discovery of cell attachment factors such as laminin, which is the surface component of the basement membrane.

    The isolation and purification of the intact laminin from normal tissue has been the most difficult task and key step to investigate the function of laminin.  I was one of those who had trouble isolating laminin from normal tissues.  Fortunately, I developed a system to isolate intact laminin from human placenta for the first time.  But 1 have not been successful in isolating laminin from other tissues yet.  From the experiences gained from human placenta, we have expanded our research into several other animals.  We have compared the nature of laminin from cow, rat, and mouse placentas.  These are similar to each other, but are distinctively different from the laminin isolated from tumors of the same species.  We would like to expand our research into an animal model system which we can apply to human diseases.  The best possible model for this is a monkey.  Preliminary data indicated that monkeys placental membranes and their laminin content are very close to that of the human.

    If we can use a monkey system to develop the experimental design, we can gain knowledge about the human system which we can not obtain from other animals.

   I have been looking for such an opportunity for quite some time.  Then, I realized that my close friend, and high school classmate, Dr. Fumiaki Cho, has been working in Tsukuba Primate Center, a new modern facility, where a large number of cynomolgus monkeys have been bred successfully.  Fortunately enough one of their research projects involved C-sections of term pregnancy.  When I went to Japan in 1984 to visit several universities to give seminars, I also visited Tsukuba Primate Center and asked if I could get permission to obtain fresh monkey placentas and process them on site.  Not only was my request granted smoothly, but.  I also got warmhearted cooperation from Director Dr. Honjo and several of his Sr. staff members.  As a result, I have obtained the necessary number of monkey placentas from which I could isolate laminin.

    This time I was asked by TPC NEWS editorial committee to write a general description of basement membrane and its component laminin, including my own area of research.

    The basement membrane is a extracellular matrix which is a necessary component of functional tissues.  These cells, namely endothelial cells and epithelial cells, must attach to the basement membrane, enabling the basement membrane to work as a selective barrier between inside of the membrane and the outside.  Blood vessels, for example, are nothing but a basement membrane structure which eliminate waste materials from tissues.

The same blood then brings in necessary nutrients, including oxygen.  Several diseases, including diabetes mellitus, cause changes in the basement membrane.  These abnormal basement membranes are thick(3-5 times of normal) and leaky to the extent that there is less barrier between the inside of the blood vessels and the outside, Large molecular weight proteins can freely leak out in such a terminal situation.  However, at this time we don't know how these changes to the membrane and cells happened.  In many tissues, including eyes, brain, kidneys, nerves, and skin, basement membranes play key roles in a variety of disease conditions, This is a reason that scientists working on basement membranes come from a variety of backgrounds, The research itself has expanded explosively in the last ten years.  We held the first international conference on basement membranes in Japan in the summer of 1985.

    The center of the basement membrane (lamina densa) contains collagen Type IV, while the outer layer (lamina rarae) is covered by laminin, proteoglycan and others.  These surface components, such as laminin and proteoglycan, also work as cell binders or as biological glue.  Many diseases such as nephritis, are actually basement membrane diseases. One of the recent developments of basement membrane research is cancer research.  In the case of metastasis, the tumor cell must attach and break through the basement membrane to get into blood vessels or lymphatic channels.  There, in order to survive, the cancer cell must again attach to the basement membrane to get out of the hostile blood stream.  The key step in metastasis is the tumor cell attachment of the basement membrane, If this mechanism were known we could control the spreading of tumor cells in our body.  Collagenase production by tumor cells, laminin receptors on the tumor cell surface have been investigated extensively.  The investigation of the tumor cell surface, and oncogene, are important but studies of basement membranes are equally important.  Basement membranes are directly involved in development and differentiation of the tissues.  However, detailed structure and function of basement membranes are not 'yet known. The majority of investigators who are working on laminin are using tumor systems, because these tumors produce abnormally large amounts of laminin, which may be defective.  These tumor laminins are soluble and easily purified. On the other hand, normal tissues contain only small amounts of laminin and this laminin is not easily soluble.  It is difficult to isolate the intact molecule from normal tissue.  Tumor laminin consists of two subunits A (400,000 MW) and B ( 200,000 MW).  Laminin isolated from normal placenta has three subunits A, B, and M ( 300,000 MW). The M is taken from the first letter of my daughter's name, Mika, who named this subunit,

    If the absence of the M subunit in tumor laminin is not due to tissue specificity, but only due to neoplastic transformation, the subunit M is a marker protein, which many scientists have been looking for.  However, subunit M has only been detected from placentas (human, cow, rat, mouse and monkey) and because other tissues are difficult to extract, the presence of laminin M has not been proven yet.

    On the other hand, we have examined only five tumors (human, rat and mouse).  Therefore, we cannot definitively conclude this difference is due only to tumor transformation.  We have just started a tumor project to elucidate or confirm our initial findings.  Monkeys will be useful in this project because tumors can now be induced in healthy animals. 

    Another interesting project is to confirm one of the hypothesis that eclampsia, sudden contraction of the uterus which leads to the death of the fetus, is due to the mother's antilaminin antibody rejecting the placenta.  The monkey is the perfect animal model. Other animals are less desirable because of differences in gestation period, placental structure and function.

    As a conclusion, I have described the importance of basement membranes and the area of my own research.  I have also mentioned how monkeys can be useful as an experimental model for us.  Even though I have never approached monkeys before, I came to the Tsukuba Primate Center.  Now, I am kind of familiar with those monkeys and monkey tissues. This familiarity is largely due to the warm cooperation given to me from the efficient professionals who are operating Tsukuba Primate Center.

    I hope that the usefulness of monkeys as a medical research model is more widely recognized among basic medical scientists. I would like to extend my appreciation to several key staff members who have been patient with me and guided me in accomplishing my initial purpose at the Tsukuba Primate Center.  Thank-you again.

 

Page 15 Overseas Topics - A side of the animal protection movements in the U.S.A.

    A short paper titled "How Radical Animal Activists Try to Mislead Humane People" has been presented in Laboratory Primate Newsletter, 23(3) '84.

    The authors, D. Caroline Coil & Neal E. Miller, have reported the result of an survey, which had been conducted to determine how true and typical the accusations being stated in a publication of the animal activists group were:

     The survey covered every article (a total of 608) appearing for the last five years in the Journals published by the American Psychological Association.

     The accusations and the percentage occurrences of articles in which such accused treatments and/or facts were reported were:

(1) "Animals are given intense, repeated electric shocks which they can not escape, until they lose the ability to even scream in pain any longer"- 0.0%.

(2) "They are deprived of food and water to suffer and die slowly from hunger and thirst"- 0.0%.

(3) "They are put in total isolation chambers until they are driven insane or even die, from despair and terror"- 0.0%.

(4) "They are subjected to crushing forces which smash their bones, and rupture their internal organs"- 0.0% (5) "Their limbs are mutilated or amputated to produce behavioral changes"- 0.0%.

(6)  "They are the victims of extreme pain and stress, inflicted upon them out of idle curiosity, in night marish experiments designed to make healthy animals psychotic"- 0.0%

        The authors have told that none of the allegations 'were found be true in any of the 608 articles. They also have demonstrated the contributions and needs of the animal experiments to the progress of science and medicine, introducing various facts and data.

    Then they have concluded as follows: Nobody wants to see animals suffer needlessly nor do they want progress toward the elimination of suffering in human to be halted. Therefore, it is essential to be totally honest in the representation of facts concerning animal welfare in research......  By suggesting the false and by suppressing the true, radical animal activists are diverting the energy and funds of humane groups away from areas where animal abuse is common (such as the abandonment of 10 million pet dogs and cats each year to die of starvation, disease, and road accidents) in order to focus upon an area in which it is rare.

   We can see a side of the relationship between animal protection movements and behaviorological studies using laboratory animals in the

 

Page-16 Case Report - Lung mite infection

    Recently, as far as our own data are concerned, the incidence of lung mite infection in imported wild cynomolgus monkeys has strikingly decreased.  From 1982 through 1984 no case of this infection has been found at the autopsies of 150 dead wild-originated monkeys, whereas the positive rate in the survey carried out in 1963 by Honjo et al, was about 22%.  The causative mite, Pneumonyssus simicola belongs to the Mesostigmata.   It usually parasitizes in lungs, whereas ordinary mites parasitize in skin and hair.  The lesion in the lung is pale yellow in color with a slight gloss and slightly upheaval, showing clear demarcation, The size of the lesion ranges from0.3 to 1.1cm with the mode of 0.3 cm(Fig. 1).  The mites are found in the yellowish pus on the cut surface of lesions.

    The pathological findings of the lesion are as follows: the round cells crowd around the mite, making circular layers.  A lot of brown pigment is observed.  Smooth muscular tissues are often recognized in the lesion (Fig. 2).  Since male and female adult mites as well as larvae are found in the lesions, this species of mite appear to live their whole life in the lung.  The morphological characteristics for the classification of this mite are the presence of the stigma with short peritrema and of the dorsal plate of a shape of shield on their back (Fig. 3).  It is thought that the lesion resulted from the movement of mites in lungs (Fig. 4), and the route of infection in monkeys might be nasal or oral.  After the infection, the mites appear to cause lesions in peripheral bronchi.

    As yet there has been no reported case of human infection by the mite species.  Judging from the infection cycle, I think, this lung mite is not so important for man from the view point of zoonosis.  As one of the reasons relating to the decrease in the incidence of lung mite infection in imported wild cynomolgus monkeys, it can be thought that Cambodian monkeys which showed high positive rate have been unable to be imported.  However, there may be some other socio-biological reasons for the decrease.

 

Page 17 A Clinical Commentary by a Veterinarian - Wounds found in a Mischievous monkey

     One morning, when I entered room no.5.  I smelled a strong odor of blood.  I thought that there might be a monkey who had wounded finger or some other body part.  I searched for the monkey.   In a group feeding cage, I found a male cynomolgus monkey aged 4 and half, whose leg was covered with semicoagulated blood.

     Immediately I and two animal technicians moved the monkey into another cage and anesthetized him. Then I carried the monkey to the treatment room and examined his wound, A horizontal slash of 5cm width and 2cm depth, as if it was cut with a sharp knife, was found at the left calf of the monkey.

    Bleeding from the end of a vein was still going on.  I first fixed forceps at the broken ends of the vein. The tips of the forceps had been covered with vinyl tubes lest they should break the blood vessels.  Then I sutured the deep part of the broken muscle to draw up both ends of the blood vessels.  Suturing small blood vessels of 1-2mm diameter without any special surgical instrument required very difficult techniques.  After suturing I took off the forceps at the cut end of the distal side then at the end of the proximal side.  Bleeding was not seen any more.  However, I had no confidence in completely maintaining the tubular structure of blood vessel. The only thing I could do was to wish that the formation of thrombus would not happen. As I was suturing, I thought that if' the vessel was obstructed, a bypassimt1g vessel would compensate for it.

I finished suturing the muscle, the fascia and finally the skin by button suture, thus I accomplished everything related to the treatment.  Then, the monkey was kept in an individual cage.  He recovered from the wound after a week.

    About one month later, the monkey was again carried into the treatment room, because he had a 5 cm long cut on his 1eft calf again.  What was the cause of these wounds?  Several days passed since he had recovered from the second injury.  While I was observing his behavior, his leg appeared suddenly from a hole of his food box.  Food boxes used in this center were fixed on the front surface of cage. The monkey put his leg through the hole to take food probably to kill time in his boring schedule.

    Wounds caused by monkey's own mischievous behavior sometimes happen, A fracture case I reported in TPC NEWS Vol.3, No.3 is an example of them.  Generally, monkeys fumble with their cages' doors, partition boards, water nozzles, keys and other things as their playthings.  They are geniuses in playing.

     By the way, the food box of the wounded monkey was taken off and his food was put directly on the cage floor.  I've never seen him in the treatment room since that time,

 

Page-18  "The Bullied" in the Monkey Colony at TPC

     Today, "bullying" is a serious problem in human society.  It is seen in the monkey society, too.

     "The bullied monkeys" in our colony are mainly one to three years old animals kept in group-feeding cages.  They correspond to from primary school pupils to senior high school students in human society.  The bullying monkeys are, of course, the far bigger ones in the cages.

    "Bullying" is done in crafty ways.  It happens, for example, when no animal technician stays in the animal room or at the time just after the lights are turned off.  The bullied can not easily take food. 

   One Sunday morning in May this year, an animal technician found a dying monkey. All of his hair had be6h fallen out: and his body was already cold. He died soon in spite of our tender care.  He was two years and two months old .

   Another case was found in June.  The bullied monkey, one year and eight months old, was the smallest in his group cage.  Fortunately, he escaped death by appropriate treatments, I think that we urgently need to take adequate countermeasures so that no more bullying occurs.