According to the present invention, there is provided a card adapter which is capable of preventing contact failure between a cover member and a ground terminal caused by the floating of the cover member when a card is inserted into a card inserting portion. A contacting surface 40 facing a back-end surface 1b of the card inserting portion 1a is provided on the ground terminal 12. A ground contacting portion 41 comprising a contacting portion 41a brought into contact with the contacting surface 40, a plate spring portion 41b extending in a widthwise direction of the card inserting portion 1a and bent in a lengthwise direction of the card inserting portion 1a, and a cantilever supporting portion 41c for supporting the plate spring portion 41b in a cantilever manner is formed in a bent portion 4a provided in a front-end portion of the cover member 4. In addition, in a lengthwise direction of the card inserting portion 1a, the contacting portion 41a of the ground contacting portion 41 is pressed against the contacting surface 40 of the ground terminal 12 to be brought into contact with each other through the elastic force of the plate spring portion 41b.

1. Field of the Invention
The present invention relates to a card adapter, connected to a ground, having a metallic cover member that covers a card inserting portion into which a card is inserted.
2. Description of the Related Art
Generally, each of card adapters typically comprise a card inserting portion, into which a card having an external terminal group exposed in a surface thereof is inserted, an adapter terminal group that is exposed in the interior and exterior of the card inserting portion and is brought into contact with the external terminal group of the card inserted into the card inserting portion, and a metallic cover member covering the card inserting portion and having a ground contacting portion which is brought into contact with a ground terminal of the adapter terminal group (for example, refer to Japanese Unexamined Patent Application Publication No. 2000-40131).
In such conventional card adapters, the cover member is arranged so as to be opposite to a bottom plate of the card inserting portion and the ground contacting portion of the cover member is brought into contact with the ground terminal in the thickness direction of the card inserting portion, to connect the cover member to the ground.
However, in the above-mentioned conventional card adapters, because the ground contacting portion of the cover member is brought into contact with the ground terminal in the thickness direction of the card inserting portion, when a card is inserted into the card inserting portion, the cover member often floats to cause contact failure between the ground contacting portion and the ground terminal. Further, as a result of having contact failure between the cover member and the ground terminal, the cover member cannot be connected to the ground. Thus, it is difficult to protect the card and the card adapter from static electricity or the like.
Accordingly, the present invention is designed to solve the above-mentioned problems, and it is an object of the present invention to provide a card adapter which is capable of preventing contact failure between a cover member and a ground terminal caused by the floating of the cover member when a card is inserted into a card inserting portion.
In order to achieve the above-mentioned object, the present invention provides a card adapter which comprises a card inserting portion into which a card having an external terminal group exposed in a surface thereof is inserted, an adapter terminal group that is exposed in the interior and exterior of the card inserting portion and is brought into contact with the external terminal group of the card inserted into the card inserting portion, and a metallic cover member covering the card inserting portion and having a ground contacting portion which is brought into contact with a ground terminal of the adapter terminal group. In addition, the ground contacting portion is brought into contact with the ground terminal in a lengthwise direction of the card inserting portion.
According to the present invention having the above-mentioned configuration, the ground contacting portion is brought into contact with the ground terminal in a lengthwise direction of the card inserting portion. For this reason, even when the cover member floats, the ground contacting portion is not separated from the ground terminal. Therefore, it is possible to prevent the contact failure between the cover member and the ground terminal caused by the floating of the cover member when the card is inserted into the card inserting portion.
In addition, according to the present invention, the ground terminal has a contacting surface facing a back-end surface of the card inserting portion, and the ground contacting portion is formed in a bent portion obtained by bending a front-end portion of the cover member. The ground contacting portion comprises a plate spring portion extending in a widthwise direction of the card inserting portion and bent in a lengthwise direction of the card inserting portion and a cantilever supporting portion for supporting the plate spring portion in a cantilever manner.
According to the present invention having the above-mentioned configuration, it is possible to press the ground contacting portion against the contacting surface of the ground terminal to be brought into contact with each other through elastic force of the plate spring portion. As a result, it is possible to stabilize the contact of the ground contacting portion to the ground terminal. In addition, since the plate spring portion of the ground contacting portion is formed so as to extend in the widthwise direction of the card inserting portion, it is possible to provide elasticity to the ground contacting portion without enlarging the size of the ground contacting portion in the thickness direction of the card inserting portion.
In addition, according to the present invention, the card adapter further comprises locking means for locking the cover member in a frame portion by sliding the cover member in a lengthwise direction of the card inserting portion in a state where the cover member is arranged at a predetermined position of the frame portion which forms sidewalls of the card inserting portion.
According to the present invention having the above-mentioned configuration, it is possible to easily assemble the cover member by the locking means.
In addition, according to the present invention, locating portions for locating the cover member in a thickness direction of the card inserting portion are formed in the cover member.
According to the present invention having the above-mentioned configuration, the cover member can be accurately arranged at a predetermined position in a thickness direction of the card inserting portion by the locating portions. As a result, the ground contacting portion can be accurately arranged toward the ground terminal.
As described above, according to the present invention, even when the cover member floats, the ground contacting portion is not separated from the ground terminal. As a result, when a card is inserted into the card inserting portion, it is possible to prevent contact failure between the cover member and the ground terminal from occurring. Therefore, it is possible to protect the card and the card adapter from static electricity or the like.

Card adapter including metallic cover member connected to ground


A dual connector system includes a host circuit board with first and second electrical connectors. The first electrical connector has a housing with a card slot. The second electrical connector has a housing having a latch member including a latching tab. The dual connector system includes a dual connector module having a module circuit board having contact pads. The dual connector module has a latching element that engages the latching tab to hold the dual connector module in the mated position with the first electrical connector and the second electrical connector. The dual connector system includes an ejector actuated to release the latching tab from the latch member to allow the dual connector module to unmate from the electrical connectors.

The subject matter herein relates generally to a mechanism for releasably securing a dual connector module in a dual connector system.
Electrical connectors are known for connecting an electronic module to a host circuit board. Such connectors are typically mounted on the host circuit board and retained on the host circuit board by soldering or some other retention feature. The electronic module must be installed in the connector and retained in the connector to provide a reliable electrical interconnection between the module and the host circuit board.
One type of electronic module includes a circuit card having a first set of contact pads along a card edge and a second set of contact pads along a bottom face of the circuit card. The first set of contacts mate with a card edge connector on the host circuit board, and the second set of contacts mate with a Z-axis connector on the host circuit board. The module is installed on the host circuit board by first moving the module downward along a Z-axis toward the host circuit board, and then translating the module horizontally along the host circuit board to install the card edge into the right angle connector. In this position the first set of contact pads are electrically connected to contacts in the right angle connector, and the second set of contact pads are electrically connected to contacts in the Z-axis connector. The module must be retained in this position to maintain the electrical connections.
There may be a need to remove the electronic module such as for replacement or upgrade. The module may be mounted in close proximity to other components on the host circuit board, thereby making it difficult to access the module and associated retention mechanism.
Accordingly, there is a need for a mechanism that securely retains an electronic module in a mated condition with an electrical connector and that can be disengaged to allow release of the module in tight spaces.


A base station equipment for receiving and transmitting a signal of a desired user, which signal to be received may arrive at the equipment along several different paths with several different delays, and which equipment includes one or more antenna arrays composed of several elements, one or more channel units having components for phasing the signal to be transmitted and received by the antenna array in such a way that the gain from the antenna array is the greatest in the desired directions. Each channel unit includes components for searching for the incoming directions and delays of the received signal components, and components for controlling the phasing components on the basis of the information in such a way that the angle of the greatest gain of the antenna beams is deflected in the environment of the desired directions.

The invention relates to a method for providing angular diversity in a base station of a cellular radio system, which system comprises in each cell at least one base station that communicates with mobile stations located within its area and that transmits a data signal to the mobile stations by using antenna beams that change in time.
The present invention is applicable for use in a data transmission system applying any multiple access method, but especially in a cellular system utilizing code division multiple access. Code division multiple access (CDMA) is a multiple access method, which is based on the spread spectrum technique and which has been applied recently in cellular radio systems, in addition to the prior FDMA and TDMA methods. CDMA has several advantages over the prior methods, for example spectral efficiency and the simplicity of frequency planning. An example of a known CDMA system is the broadband cellular radio standard EIA/TIA IS-95.
In the CDMA method, the narrow-band data signal of the user is multiplied to a relatively wide band by a spreading code having a considerably broader band than the data signal. In known test systems, bandwidths such as 1.25 MHz, 10 MHz and 25 MHz have been used. In connection with multiplying, the data signal spreads to the entire band to be used. All users transmit by using the same frequency band simultaneously. A separate spreading code is used over each connection between a base station and a mobile station, and the signals of the different users can be distinguished from one another in the receivers on the basis of the spreading code of each user.
Matched filters provided in the receivers are synchronized with a desired signal, which they recognize on the basis of a spreading code. The data signal is restored in the receiver to the original band by multiplying it again by the same spreading code that was used during the transmission. Signals multiplied by some other spreading code do not correlate in an ideal case and are not restored to the narrow band. They appear thus as noise with respect to the desired signal. The spreading codes of the system are preferably selected in such a way that they are mutually orthogonal, i.e. they do not correlate with each other.
In a typical mobile phone environment, the signals between a base station and a mobile station propagate along several paths between the transmitter and the receiver. This multipath propagation is mainly due to the reflections of the signal from the surrounding surfaces. Signals which have propagated along different paths arrive at the receiver at different times due to their different transmission delays. In the CDMA, the multipath propagation can be exploited in the reception of the signal in the same way as diversity. The receiver generally utilized in a CDMA system is a multibranch receiver structure where each branch is synchronized with a signal component which has propagated along an individual path. Each branch is an independent receiver element, the function of which is to compose and demodulate one received signal component. In a conventional CDMA receiver, the signals of the different receiver elements are combined advantageously, either coherently or incoherently, whereby a signal of good quality is achieved. CDMA systems can also apply a soft handover wherein a mobile station may simultaneously communicate with several base stations by utilizing macrodiversity.
The connection quality of the mobile station thus remains high during the handover and the user does not notice a break in the connection.
Interference caused by other connections in the desired connection thus appears in the receiver as noise that is evenly distributed. This is also true when a signal is examined in an angular domain according to the incoming directions of the signals detected in the receivers. The interference caused by the other connections in the desired connection thus also appears in the receiver as distributed in the angular domain, i.e. the interference is rather evenly distributed into the different incoming directions.
The capacity of the CDMA, which can be measured by means of spectral efficiency, has been further improved with sectorization. A cell is then divided into sectors of a desired size that are serviced by directional antennas. The mutual noise level caused by the mobile stations can thus be reduced significantly in the base station receiver. This is based on the fact that on average the interference is evenly distributed between the different incoming directions, the number of which can thus be reduced by means of sectorization. The sectorization can naturally be implemented in both transmission directions. The advantage provided in the capacity by the sectorization is proportional to the number of the sectors.
A sectorized cell may also utilize a special form of soft handover, softer handover, wherein a mobile station performs a handover from one sector to another by communicating simultaneously with both sectors. Even though soft handover improves the connection quality and sectorization increases the system capacity, the movement of the mobile stations naturally leads to the stations performing several handovers from one sector to another. This loads the processing capacity of the base station controller. Several soft handovers also produce a situation where several mobile stations communicate simultaneously with more than one (usually two) sector, whereby the increased capacity provided by the sectorization is lost as a signal of a mobile station is audible in a wide sector.
The multiple access interference of the CDMA systems has also been reduced by means of different known multiple access interference cancellation (IC) methods and multi-user detection (MUD). These methods are best suited for reducing the interference produced within the user's own cell, and the system capacity can thus be increased to a double compared to a system implemented without interference cancellation. However, these methods do not improve the size of the coverage area of the base station, i.e. the cell size does not change when these methods are used. Also, the IC/MUD techniques are complicated to realize, wherefore they have only been developed in the uplink direction, and the opposite transmission direction is similar as in a conventional CDMA system.
Another method that has been developed is an SDMA (Space Division Multiple Access) method wherein the users are distinguished from one another on the basis of their location. This is performed in such a way that the beams of the receiver antennas at the base station are adjusted to the desired directions according to the location of the mobile stations. For this purpose, the system uses adaptive antenna arrays, i.e. phased antennas, and the processing of the received signal, by means of which the mobile stations are tracked.
The use of the SDMA in connection with the CDMA provides several advantages over the prior methods, such as sectorization. If the sector beams in the sectorization are narrowed in order to increase the spectral efficiency, the number of the handovers to be performed from one sector to another also increases. This in turn increases too much the calculation capacity required in the base station controller.
In connection with the application of the SDMA, the background art is illustrated in A. F. Naguib, A. Paulraj: Performance of CDMA Cellular Networks With Base-Station Antenna Arrays (Proc. International Z urich Seminar on Digital Communications, pp. 87-100, Zurich, Switzerland, March 1994), which is incorporated herein by reference. In the SDMA a signal is thus received by means of an antenna array, and the received signal is shaped by means of digital signal processing in such a way that the directivity patterns of the antennas are suitable for the stages following the shaping in the receiver. In prior art arrangements, the received signal is shaped in order to maximize the signal-to-interference ratio of the desired signal. The received signal is thus shaped in such a way that the directivity pattern of the antenna array minimizes the interference caused by the other connections in the desired signal. In the arrangement according to the aforementioned reference, each detected signal component is subjected to individual beam shaping, i.e. the impulse response must be known before the shaping.
Experimental Studies of Space-Division-Multiple-Access Schemes for Spectral Efficient Wireless Communications by G. Xu, H. Liu, W. J. Vogel, H. P. Lin, S. S. Jeng and G. W. Torrence (IEEE Int. Conf. On Comm. ICC 1994, New Orleans, USA, IEEE 1994), which is incorporated wherein by reference, discloses a method which applies the SDMA and in which the directivity pattern of the transmitter antennas is shaped. However, the method disclosed is suitable for use only in systems where both transmission directions are on the same frequency.

Method for providing angular diversity, and base station equipment


A wedge-shaped section is located near the top, near portion of a trailer hitch insert tube. The wedge-shaped segment is located opposite a complementary shaped section of the insert tube. The wedge-shaped section engages the insert tube to reduce or eliminate movement of a trailer hitch insert tube within a trailer hitch tube attached to a vehicle.

There are a variety of trailer hitches and towing attachments in use in the marketplace today. A ball-type trailer hitch is fairly common and can be either permanently or temporarily-attached to the frame or bumper of a vehicle. Many boat trailers, camping trailers, and moving trailers employ a ball-type attachment.
A ball-type trailer hitch can be permanently attached to a motor vehicle by welding or bolting a tongue to the underside or frame of a motor vehicle, and mounting a 2-inch ball on the portion of the tongue that extends beyond the vehicle.
Alternatively, a ball-type trailer hitch may also be permanently attached to a vehicle by bolting or welding hollow tubing to the underside of a motor vehicle. Typically, a tube having inner dimensions of approximately 11/2 or 2 inches by 11/2-2 inches protrudes from beneath a vehicle. A second tube, having outer dimensions slightly smaller than the opening in the tube attached to the vehicle is inserted into the tube attached to the vehicle. A steel pin, or other suitable fastener, is fed through both sets of steel tubes and serves to hold the inserted tube in place. The pin is typically 1/2-1 inch in diameter. A ball-type trailer hitch is affixed to the protruding end of the inserted steel tubing, and provides a point of attachment for a trailer.
This type of trailer hitch is typically rated at about 500 lb. tongue weight, and a 10,000 lb. load.
The tube-type trailer hitch attachment has also been used to mount bicycle racks, motorcycle racks, and other carrying devices to a vehicle. The tube-type hitch attachment has become more popular in recent years because of the versatility in allowing a vehicle owner to install a single-style hitch which can accommodate a variety of carrying or towing needs. One disadvantage of the tube-type hitch attachment is the movement and consequent noise caused by the inserted tube shaking or vibrating within the surrounding tube.
In order to allow the inserted tube to be readily removed from the hitch assembly, it is necessary that the inserted tube be sufficiently smaller than the surrounding hitch tube. Otherwise, frictional forces acting upon the inner tube restrict its removal and thereby increases the burden of disengaging the towing assembly. Conversely, where the inner tube is sized to decrease the frictional force so that removal is facilitated, the inner tube may bounce within the outer tube thereby creating oscillation and noise or rattle. This rattle can be extremely distracting to the driver of the vehicle, especially if the driver is not familiar with the movement and noise commonly made by trailer hitches.
The movement and consequent rattle can cause the driver to pay excessive attention to the trailer, or other carrying apparatus inserted in the hitch tube. This distraction serves as both an annoyance and a danger to the driver, as the driver's full attention is no longer devoted to other vehicular traffic, road conditions, and pedestrian traffic. In fact, the rougher the road conditions, the more trailer hitch will tend to move and rattle because of the inertia developed by the apparatus attached to the inserted tube.
A need therefore exists for a trailer hitch insert tube which minimizes or eliminates movement and noise generated by the trailer hitch.


A method of radiation-free hematopoietic stem cell (HSC) transplantation comprises administering to a mammalian subject one or two doses of 2 to 10 mg/kg body weight of a purine base analog, such as 6TG as a pre-conditioning step. The method further comprises engrafting into the subject hypoxanthine-guanine phosphoribosyltransferase (HPRT)-deficient donor HSCs within 48 to 72 hours of the pre-conditioning step; and administering to the subject about 1 to 5 mg/kg of the purine base analog every two to four days for two to eight weeks following the engrafting step. The method is performed in the absence of pre-conditioning via radiation. The subject is therefore not treated with myeloablative radiation in preparation for transplantation, and thus the subject is free of myeloablative radiation-induced toxicity.

Hematopoietic stem cell transplantation (HSCT) is a mainstay of treatment for many hereditary disorders and lymphohematopoietic malignancies (1). Furthermore, hematopoietic stem cells (HSC) in general represent an important target for ex vivo gene therapy. Gene transfer into HSC provides a potential strategy for both correcting monogenic defects and altering drug sensitivity of normal BM to cytotoxic drugs. These applications have significant therapeutic potential but have been limited by low gene transfer into HSC. Recent advances, such as improved cytokines for minimizing commitment during ex vivo manipulation, fibronectin-assisted gene transfer and enrichment of HSC prior gene transfer have improved the efficiency of gene transfer into human cells and enhanced human gene therapy trials (2). However, the efficiency of gene transfer into HSC and the engraftment of large numbers of transduced cells remains a major challenge to broaden the application of this technology for the successful treatment of cancer and monogenetic diseases.
In order to enhance engraftment of gene-modified HSC and to decrease the time needed for lymphohematopoietic reconstitution after HSCT, in vivo selection strategies employing drug resistance genes such as dihydrofolate reductase (DHFR) (3) or multiple drug-resistance gene 1 (MDR1) (4, US1996/017660) have been tested but have generally failed due to unacceptable toxicity (5) or insufficient selection efficiency (6). Currently, mutant forms of O6-methylguanine-DNA-methyltransferase (MGMT) are being tested for their ability to confer chemoprotection against BCNU or temozolomide in combination with O6-benzylguanine (2,7, US1997/004917), but these agents pose a considerable risk of toxicity, and recent observations suggest that mutant MGMT may confer a selective disadvantage when expressed at high levels (8). In US2003032003AA a selection strategy has been described for selecting HPRT-deficient cells in vivo by 6TG. However, in this patent application either irradiation is still used for preconditioning prior in vivo selection or in vivo selection is performed in cycles with recovery periods in between 6TG administration.
Furthermore, the suggested 6TG dose is high and administrated over a long time period (200 mg/kg total dose over 55 days). In addition, an approach to inactivate HPRT expression in BM and subsequently select the donor cells with 6TG in vivo has been reported by Porter and DiGregori as "interfering RNA mediated purine analog resistance" ('iPAR'). This report demonstrated the feasibility of HPRT inactivation in HSC with a lentiviral vector expressing shRNA targeting Hprt and enrichment for these altered hematopoietic cells with 6TG in mice in vivo. However, in this report, pre-conditioning was still performed by total body irradiation, and in vivo chemoselection was not initiated until at least 4 weeks post-transplant. In addition, 6TG was administered either as a short pulse or at dosages chosen to be only moderately myelosuppressive, and it is not clear whether adequate levels of HSC transduction were achieved by the second-generation lentiviral vectors employed in their study. Overall, the engraftment levels reported were variable and relatively modest.
There remains a need for more effective methods of HSCT that avoid toxicity while reconstituting bone marrow cells.

This patent is subject to a terminal disclaimer.

THE REGENTS OF THE UNIVERSITY OF CALIFORNIA


A radio network provides centralized load control in order to control uplink interference in a cell while at the same time permitting adaptation to fast channel variations. The radio network sends to a mobile terminal a percentage of time that the mobile terminal may transmit over a radio channel in the uplink direction to the radio network. The mobile determines a current or future condition of the radio channel and restricts uplink transmissions based on the received percentage and the radio channel condition. Transmissions are restricted to times when the current or future condition is favorable up to the percentage amount. Channel condition favorability may be determined by the mobile station, in one example embodiment, by comparing the radio channel condition to an average condition of the channel, such as a local or moving average channel condition. Also in an example embodiment, the mobile may determine a current or future channel condition using transmit power control commands received from the radio network.

In cellular communications systems, the mobile radio station communicates over an assigned radio channel with a radio base station. Several base stations are coupled to a switching node which is typically connected to a gateway that interfaces the cellular communications system with other communication systems. A call placed from an external network to a mobile station is directed to the gateway, and from the gateway through one or more switching nodes to a base station which serves the called mobile station. The base station pages the called mobile station and establishes a radio communications channel. A call originated by the mobile station follows a similar path in the opposite direction.
Due to the rapid expansion of wireless mobile communications and the need for wideband multimedia services, there is a continuing need to better utilize the available frequency bandwidth. A common strategy in Frequency Division Multiple Access (FDMA)/Time Division Multiple Access (TDMA) systems is to reuse the frequencies in the network. The challenge with frequency reuse is to counteract or at least reduce the interference between transmitters in the system using the same frequency by controlling the transmit power levels of the radio signals and by separating to the extent practical the transmitters by a sufficient geographic distance. The transmit power levels of the mobile stations and base stations are ideally lowered so that only the minimum transmission power necessary to maintain satisfactory call quality is used. By reducing mobile and base station transmission power, the other radio communicators experience lower interference which means that the system capacity may be increased.
In a Code Division Multiple Access (CDMA) mobile communication system, spreading codes are used to distinguish information associated with different mobile stations or base stations transmitting over the same radio frequency band—hence the term "spread spectrum." In other words, individual radio "channels" are discriminated upon the basis of these codes. Because all users of a CDMA communications system transmit information using the same frequency band at the same time, each user's communication interferes with the communications of the other users. In addition, signals received by a base station from a mobile station close to the base station are much stronger than signals received from other mobile stations located at the base station's cell boundary. As a result, distant mobile communications may be overshadowed and dominated by close-in mobile stations.
Interference is a particularly severe problem in CDMA systems. If one mobile station transmits at a power output that is too large, the interference it creates degrades the signal-to-interference ratio (SIR) of signals received from other mobile radios to the point that a receiving base station cannot correctly demodulate transmissions from the other mobile radios. In fact, if a mobile station transmits a signal at twice the power level needed for the signal to be accurately received at the base station receiver, that mobile's signal occupies roughly twice the system capacity as it would if the signal were transmitted at the optimum power level. Unregulated, it is not uncommon for a "strong" mobile station to transmit signals that are received at the base station at many, many times the strength of other mobile transmissions. Such a loss of system capacity to excessively "strong" mobile stations is unacceptable.
Additional problems are associated with excessive transmit power. One is the so-called "party effect." If a mobile transmits at too high of a power level, the other mobiles may increase their respective power levels so that they can "be heard," compounding the already serious interference problem. Another problem is wasted battery power. It is very important to conserve the limited battery life in mobile radios. The major drain on a mobile's battery occurs during transmission. A significant objective for any power control approach, therefore, is to reduce transmit power where possible.
Most radio transmit power control procedures try to keep the signal strength and/or quality of the signal detected by a receiver above a desired threshold without using unnecessarily high transmit power. Because power control is so important, CDMA systems employ a relatively high sampling rate for the power control algorithm, e.g., 1500 times per second. To minimize overhead control signaling, only one bit is used to communicate power control adjustments to the radio transmitter. The power is stepwise increased or decreased based upon a comparison of received signal strength, or some other signal parameter, with a threshold. The receiver controls the transmitter's power by issuing transmit power control commands (TPCCs)—power up or power down—at the same high sampling rate (e.g., once every 0.667 msec) based on signal quality measurements, e.g., signal-to-interference ratio (SIR). If the measured signal parameter value is less than a target signal parameter value, the power-up command is issued; otherwise, the power-down command is issued. The radio transmitter responds to the power control commands by increasing or decreasing its transmit output power level P, for example, by a certain incremental power step Δ, i.e., P←P+Δ or P←P−Δ.
Because a power-up or a power-down command issues every 0.667 msec, the transmit power level is never constant or static. Accordingly, even in an ideal radio environment, the incremental power control commands continually alternate between power up and power down so that the transmit power level and the received signal quality oscillate up and down an incremental step around a target value. In order to maintain the quality of the received signal always above a prescribed limit, the target value needs to be set slightly higher than that limit so that the received signal quality after the power down step is still above the prescribed limit.
Uplink capacity is limited by a maximum, acceptable level of uplink interference. Uplink interference depends on the current uplink traffic load, movement of the mobile station (mobility), and current radio conditions. Traffic load can vary considerably and rapidly, especially with multimedia and other "bursty" data services. Mobility and changing radiowave propagation conditions affect the quality of communications channels, and thus, the proper power level needed to transmit over those channels.
FIG. 1 shows a mobile communications environment with multiple, direct and indirect radio signals between base stations and mobile stations. FIG. 2 illustrates that the overall signal strength of a received signal varies as the distance between the transmitter and the receiver and depends (in one well-established model) on three variable factors: path loss, shadowing, and multipath fading. Path loss is the overall decrease in the field strength of the transmitted signal as the distance between the transmitter and the receiver increases. Shadowing occurs as a result of obstructions between the transmitter and receiver such as buildings, trees, and other objects in the environment. Multipath fading occurs as a result of constructive and destructive interference between multiple waves (rays) reaching the receiver from the transmitter. Multipath fading is particularly troubling because the quality of the received signal varies so rapidly.
In light of these obstacles, there is a need to dynamically control uplink interference so that satisfactory service is provided and maximum capacity is achieved. One way to control uplink interference is to assign maximum bit rates to mobile transmitters based on the current uplink interference load. But maximum bit rates do not adapt to changing conditions. Moreover, when most or all mobiles are transmitting at their maximum bit rate, there will be a spike in the uplink load perhaps causing less than satisfactory service. At other times, many of the mobiles may not transmitting at all, or at less than maximum bit rate, causing a dip in the uplink load and unused capacity.
Another approach is to limit the amount of time that mobiles can transmit in the uplink direction. A mobile station transmitting only 50% of the time generates less interference than when transmitting 100% of the time. One way to implement time-limited transmission is to assign each mobile station a probability of transmission and have each mobile transmit in a random fashion to meet this probability. EP 1033846 A1 describes broadcasting access probabilities to mobiles. Each mobile compares a random number to the probability. Uplink transmission occurs only when the random number is less than the broadcast probability.
Although this approach statistically reduces the uplink load, its randomness means that there is no attempt to transmit at particular times and not at others. As a result, the random time chosen to transmit may very well correspond to a time when radio transmission conditions are unfavorable, e.g., during a fading dip or when the current uplink load is momentarily high. An unfavorable condition means that the mobile must increase its transmit power, if possible, causing increased battery drain and interference or decreased signal quality at the base station.
These problems are overcome by limiting the amount of time that a mobile station can transmit and timing transmissions so they occur during favorable channel conditions rather than during less favorable channel conditions. Transmitting during favorable channel conditions requires less power, which translates into lower battery consumption and less interference to other uplink transmissions. Transmitting during favorable channel conditions also means fewer bit errors in received transmissions. Less uplink interference translates into a reduced uplink traffic load and more uplink capacity. If desired, the additional capacity may be used to allow mobile terminals to transmit at higher bit rates than would otherwise be permitted/possible in unfavorable channel conditions.
Having the mobile terminals transmitting during favorable channel conditions is particularly beneficial because fast channel variations due to constructive and destructive multipath fading are typically uncorrelated between mobile stations. If mobile stations only transmit during constructive fades (rather than during destructive fades), a higher instantaneous bit rate can be used to maintain the same average bit rate as when transmitting continuously. Because fading is uncorrelated between the transmitting mobiles, the number of simultaneously transmitting mobile stations will, on average, decrease. In this way, interference is reduced.
The radio network provides centralized load control in order to control and limit uplink interference in a cell while at the same time permitting adaptation to fast channel variations. The radio network sends to a mobile terminal a percentage of time that that mobile terminal may transmit over a radio channel in the uplink direction to the radio network. The mobile determines a current or future condition of the radio channel and restricts uplink transmissions based on the received percentage and the radio channel condition. Transmissions are restricted to times when the current or future condition is favorable, up to the percentage amount. Favorability may be determined by the mobile station, in one example embodiment, by comparing the radio channel condition to an average condition of the channel, such as a local or moving average channel condition.
Based on the load for the overall cell, an activity factor corresponding to a fraction or percentage is determined for each of the mobile terminals. The activity factor defines an amount of time relative to a total time that the mobile terminal may transmit based upon desired uplink load level for the cell area. The activity factors need only be sent to the mobile terminals infrequently thereby keeping the signaling load low. Activity factors could be sent more frequently, if desired, depending upon changing load conditions. Moreover, one activity factor may be assigned to a group of mobiles or to all mobiles in a specific cell, group of cells, or even in an entire radio network. Broadcasting group-specific, cell-specific, and network-specific activity factors reduces signaling. In any event, the mobile terminals are permitted to implement an appropriate algorithm to determine the best times to transmit when favorable channel conditions are present for that particular mobile terminal.
Information regarding the current or future condition of the radio channel is provided, in an example, non-limiting embodiment, from the radio network, preferably at a high frequency. Because transmit power control commands (TPCCs) are sent from the radio network very frequently already, TPCCs are advantageously used by the mobile terminals, in a preferred example embodiment, to determine when favorable channel conditions exist or will exist. One of the benefits of using TPCCs is that the radio network can control the uplink load without having to send additional control signaling messages to each of the mobile terminals to indicate when each mobile terminal should transmit and when it should stop transmitting.
One example algorithm that uses TPCCs to determine channel conditions calculates a cumulative transmit power control command (CPTCC) value. The network provides an activity factor used to determine a transmit threshold. The cumulative TPCC value is compared to the transmit threshold, and the determination of when to transmit is based on the comparison. When the percentage of allowed transmit time is higher, the transmit threshold is higher, and when the percentage is lower, the transmit threshold is lower.
Because transmissions occur during favorable channel conditions when there is less interference and because those favorable channel conditions are not correlated between different channels, the overall uplink cell interference level decreases. This permits the mobiles to increase their bit rate up to a preset maximum, if desired, or conserve battery power. If the interference level is reduced, the cell capacity increases. These advantages are not achieved when transmissions are timed randomly.

Method and apparatus for uplink transmission timing in a mobile communications system


A method for leak testing of hand-held piston stroke pipettes and a corresponding leak testing apparatus (1) for performing the method. Essential to the system is a holder (8) for the sealed connection of a pipette tip (3) to the leak-testing apparatus. A line system (9) with connecting sections of the leak-testing apparatus forms a specific connection volume. The pipette tip (3) is evacuated to a measuring pressure below atmospheric pressure. With the vacuum pump (5) disconnected and the connection volume sealed off, the pressure is measured after a specific time interval, and any possible rise in pressure is determined. An indication of the serviceability of the pipette (2) is generated from the pressure rise determined.

1. Field of the Invention
The invention relates to a method of leak testing of hand-held, single-channel or multichannel piston stroke pipettes, preferably of those with air cushions. The subject matter of the invention is also a leak testing apparatus which is suitable for carrying out this method. The subject matter of the invention is finally a system which includes the corresponding leak testing apparatus.
2. Description of Related Art
Piston stroke pipettes are likewise known as hand-held single channel measurement devices (German Patent Application DE 39 03 241 A1) as well as multichannel measurement devices (German Patent Application DE 100 13 511 A1 and corresponding U.S. Pat. No. 6,627,160 B2). In the process of aspiration of a sample liquid, negative pressure is produced in the piston stroke pipette and in the pipette tip which is slipped onto a slip-on section of the pipette. Sample liquid is sucked into the pipette tip by this negative pressure. Piston stroke pipettes, as direct displacers, have no air volume or only a minimum air volume above the liquid level in the pipette tip. They are less suited for biological samples since, there, it is necessary to work largely free of contamination. Therefore, air cushion piston stroke pipettes are more common in which there is a distance between the bottom of the piston and the liquid level such that the liquid level is always only in the interchangeable pipette tip which is generally made as a disposable part, while the piston bottom moves distinctly above the liquid level and thus is protected against contamination.
In piston stroke pipettes and especially in air cushion piston stroke pipettes, the requirement of tightness of all connecting sites of the pipette and the pipette tip is very strict. Due to the differential pressure during suction, air can unintentionally flow into the dead space above the liquid level via a leak. The volume which has been enlarged by the piston stroke then no longer agrees with the actually changed volume.
The sealing sites of piston stroke pipettes are subject to wear during operation. Ageing, chemical attack and other ambient influences have adverse effects, even for static seals. One especially sensitive sealing site, in addition to the dynamic piston seal, is the connection between the pipette tip and pipette on the slip-on section. For hand-held piston stroke pipettes, the process of slipping on the pipette tip takes place manually and essentially free-handed. The surfaces of the slip-on section and pipette tip are highly stressed in doing so. Moreover, new pipette tips are always being used (disposal part) so that the production tolerances of the pipette tips and the surface quality of the sealing surfaces of the pipette tips become noticeable. As a result, after longer use ridges form on the slip-on section of the pipette which can ultimately lead to a leak. Therefore, a slowly rising leakiness is characteristic for piston stroke pipettes.
Not every leak leads to a loss of serviceability of a piston stroke pipette. A very small leak can be tolerated. If the leak has only a very small flow cross section, air flows only very slowly into the dead volume of the pipette and the leak can still be within the framework of the acceptable fault tolerance.
For piston stroke pipettes, it is thus part of normal laboratory operation that the pipettes are subjected to leak testing from time to time. This is regardless of whether they are hand-held piston stroke pipettes or automatic pipetting devices. Even in the latter, there is still a corresponding risk of leaks, all the more considering that observation is limited or not possible at all.
For automatic pipetting devices, in the past, only an optical leak test was performed and such testing was only able to identify coarse leaks. According to this traditional method, the operation of the automatic pipetting device is temporarily stopped after the pipette tip into which the sample liquid has been sucked has been moved upward from the liquid level. An operator visually checks whether sample liquid is dripping from the pipette tip. For this method, a highly experienced operator is required. Moreover, this method can only be used for large leaks and large amounts of liquid. For small volumes of sample liquid, for example, of 10 μl, this method fails.
Therefore, for an automatic pipetting device, an automated method for leak testing has already been proposed (German Patent DE 693 26 773 T2) in which the pressure which is prevailing in the pipetting device, therefore in the dead volume of the pipette and pipette tip, is measured and evaluated in different pipetting states. For this purpose, the pipetting device is itself provided with a pressure sensor which detects the pressure in the dead volume between the liquid level and the piston bottom of the piston pump of the pipette. An electronic control apparatus compares the internal pressure which has been detected by the pressure sensor after taking in of the sample liquid relative to the atmospheric pressure of the ambient atmosphere. In this way, a major leak can be ascertained which makes aspiration of the sample liquid more or less completely impossible. In a second step, the internal pressure with the piston pump stopped after a certain time interval is measured and compared to the internal pressure which prevailed immediately after the end of aspiration. Depending on the pressure rise which has been determined in this method, a leak warning for the automatic pipetting device is delivered or the automatic pipetting device is released for further precision pipetting.
In the above explained known method for leak testing, in an automatic pipetting device, typical volumes of the sample liquid taken in during the leak test are 200 to 500 μl distilled water. The time interval during which pressure in the dead volume of the pipette is measured is typically 30 s compared to the first measurement immediately after stopping the pump after roughly 500 ms. This patent does not disclose anything about pressure conditions.
A method for leak testing in automatic pipetting devices cannot be applied to hand-held piston stroke pipettes, preferably air cushion piston stroke pipettes. To do this, the pipette itself would have to be equipped with a corresponding leak testing apparatus. This cannot be justified in terms of costs and would not be commercially acceptable. For this reason, manual methods for leak testing which can be carried out externally are used for hand-held piston stroke pipettes.
In a known method (Eppendorf, Pipettentester 4708), a bottle which stores a liquid, a filled hose connected to it, and an adapter are used. The adapter is connected to the attached pipette tip. The pipette arranged with the pipette tip up. The liquid level which is settling is marked. After several actuations of the piston of the pipette the position of the liquid level is checked again, the pipette having to be held exactly at the same height. If the liquid level has been established at a lower level, the system of pipette and pipette tip is leaky.
Another similar method is also known in which a hose which leads to a leak testing apparatus is connected to the pipette tip which has been slipped onto the pipette. The leak testing apparatus has a hand-actuated piston pump which produces a negative pressure and an analogously operating manometer which is connected to the piston pump on the intake side as a pressure sensor. The vacuum pump is actuated by hand until a sufficient measurement pressure below atmospheric pressure is established which is shown on the manometer. In this position, it is attempted to fix the vacuum pump by hand so that its pump volume no longer changes. Then, the pipette is repeatedly actuated with the other hand. If afterwards the manometer shows a higher pressure than previously, a leak in the system of pipette and pipette tip must be concluded to exist.
The method for leak testing of hand-held, single-channel or multichannel piston stroke pipettes, preferably of those with air cushions, which is carried out with the above explained device constitutes the point of departure for the teaching of this application.
In addition to the above explained known method, a method is known which works with a special measurement pipette (German Utility Model DE 82 30 186 U1). Here, a measurement pipette is connected to the pipette via an adapter. The sample liquid is taken up directly by the measurement pipette. The taken-up volume can be read directly on its scale. This volume can be compared to the setpoint which has been set on the pipette. In this method, it is disadvantageous that the air cushion, therefore the dead volume, does not agree with the air cushion which is actually present in the pipette in normal operation. The hydrostatic pressure of the taken-up liquid is also greater. Consequently, the method with use of the measurement pipette does not correspond to the actual method in working with the pipette itself.
Studies have shown that different parameters arise in pipetting with different pipettes and pipette tips which influence the results of leak testing in wide limits. The operator must ascertain by experience which parameters must be maintained and to what extent. Such a parameter is the minimum negative pressure below atmospheric pressure which forms in the intake process (see the explanation of German Patent Application DE 39 03 241 A1 above). The viscosity of the sample liquid, the strength of the reset spring which acts on the piston of the piston pump and the flow cross section of the pipette tip influence the negative pressure which is being established. The pipette should be tested at least with this pressure in order to correctly detect the amount of gas flowing in at the leak and in a manner suitable for practice. Another parameter is the pressure rise over a certain time interval. It is dependent on the size of the dead volume in the unit consisting of the pipette and pipette tip during the leak test. Even the initially explained method for leak testing in an automatic pipetting device does not take this quantity into account. Consequently a lowermost boundary value must be stipulated itself for the sake of reliability. The result is that pipettes which are in fact still usable are also serviced.

Method for leak-testing hand-held piston stroke pipettes and associated leak-testing apparatus


A disaster tolerant computer system for protection of electronic databases is achieved by cascading storage controllers to accomplish local and remote disk shadowing. A host computer is connected to a local superordinate storage controller via a local data bus. The superordinate storage controller transmits data via a long haul data link to a remote subordinate storage controller. Additionally, the superordinate controller transmits data to a local subordinate storage controller via a local data bus. Consequently, in-band storage capacity is provided remotely, as well as locally, to increase data storage capacity for database shadowing. Furthermore, the cascading of controllers may be used to increase the effective storage capacity of the computer system by using the increased storage capacity for the writing of non-redundant data.

Computer systems have become the primary means for individuals and organizations to manipulate and store data. Typically, the larger the amount of data to be stored and manipulated, the larger the reliance on a computer system. Often, large amounts of data are stored in one or more databases, where the data is organized into a database structure. Today, it is increasingly common for an entity which is responsible for controlling significant amounts of data to institute computer based systems, using electronic storage of data and databases. For example, banks, insurance companies, investment companies, retailers, government agencies, universities, and others all keep databases of customer lists, accounts, inventories, and so on in electronic storage.
When information in a database is considered critical, it is often considered prudent to take steps to preserve such data. A common approach to preserving electronic data is to use a computer system which not only stores the new data in databases, but also makes copies of those databases. In one approach, database copies are made at regular fixed intervals of time, e.g., a daily copy of a stores sales data for the day. Periodic copies, also referred to as back-ups, are used when the risk of loss of information is not so great as to justify the additional expense associated with real-time copies. Using another approach, copies may be made in "real-time", where the copied data is kept current by copying the main database as it changes. One example where such real-time copies are customary is in the banking industry, where large banks may have thousands of accounts changing each minute and the loss of the data for the myriad transactions throughout the day could prove catastrophic. The process of writing data or databases in real-time to multiple storage locations is referred to as "shadowing" or "mirroring", which yields multiple copies of the data or database.
With the need to keep multiple copies of such large amounts of data comes the need for large data storage capacities. The storage of large amounts of data is typically referred to as "mass storage", wherein the data is stored in mass storage devices. With large computer systems and networks serving an increasingly wide variety of entities and storing a wide variety and amount of data, mass storage devices are becoming the norm. One increasingly popular form of mass storage device is a disk array. A disk array is commonly defined as a collection of disks combined, coordinated and managed by some form of array management software. An increasingly popular form of disk array is the "redundant array of independent disks" (RAID). A RAID is a special type of disk array which is intended for uses involving the storage of redundant, i.e., copied, data.
Depending on how a RAID is configured and used, it can be classified as being of a certain RAID Level, i.e., Level 0-6, defined by the RAID Advisory Board, Inc. (RAB). The RAB is an organization which seeks to standardize and promote the use of RAID devices and related products. The RAB defines RAID levels in their publication The RAIDBook, Nov. 18, 1993.
There are various methods of storing information in a RAID, e.g., shadowing or striping data. When data is written to a storage device by striping, it is broken into blocks of data, wherein the blocks of data are written, or distributed, among the disks within the array. Disk striping is used to expand the overall storage capacity of a computer system for the storage of original, i.e. non-redundant, data. With disk striping, copies of original data are not made. Therefore, strictly speaking, a disk array configured for disk striping is not a true RAID. However, within the art, disk striping may be referred to, and is defined by the RAB, as a "RAID Level 0" system. As an example, a disk array which comprises six one-gigabyte disks is capable of storing about six gigabytes of new data. However, none of the newly stored data is also written to another location within the disk array, to yield a copy. The disk striping system of this example may be said to have an "effective storage capacity" of six gigabytes. Effective storage capacity refers to how much non-redundant data can be stored by a system or disk array.
Within RAID Levels 1-6, there are a variety of configurations which all store redundant data in slightly different ways. For illustrative purposes, the writing of redundant data by disk shadowing will be emphasized. Basically, disk shadowing is the creating of copies of data by writing the data to multiple disks within an array in real-time. Therefore, shadowing yields less effective storage capacity than does disk striping. For example, if there was a RAID of six one-gigabyte disks configured for disk shadowing, the effective storage capacity of the RAID may be calculated to be one gigabyte, assuming five of the one-gigabyte disk drives are used for storing redundant data.
Another characteristic of the current information age is the increasingly distributed nature of computer systems. In such distributed systems, it is often the case that access to a main database may be gained from a wide variety of remote locations. For example, an insurance company may have satellite offices throughout the country, each with a local area network (LAN) which connects to a remote central computer network over a long haul link, wherein the central network includes devices which store the company's databases of client policy information. Computers in the satellite offices rely on connectivity to the remote central computer network for access to client policy information. However, if the remote central computer network losses connectivity with the satellite offices by, for example, a power outage or equipment failure, the overall system becomes inoperable. In another example of a distributed computer system, a bank may use centralized databases, with certain degrees of access to client accounts available at automated teller machines (ATMs) worldwide. As with the insurance company example, loss of connectivity to the bank's central databases, or their corruption or malfunction, causes the entire system to become inoperable. With such distributed computer systems and reliance on critical data contained therein, it is desirable to achieve the benefits of database copies within the distributed system.
As shown in FIG. 1, a computer system using typical prior art devices is capable of storing and copying data. The host computer 110, may be a workstation, such as an Alpha workstation by Digital Equipment Corporation ("Digital") of Maynard, Mass., or a server on a LAN. The host computer 110 runs applications which seek to manipulate or store data using the disk controllers 120 and 140. The host computer 110 communicates with controller 120 via a local data bus 115. The local controller 120 interfaces with the local data bus 115 at what is referred to as its "front-end". The front-end of the controller 120 receives data to be written to storage devices in connection with the controller's 120 "back-end". The local controller 120 channels the data provided by host computer 110 to a number of devices at its back-end over buses 130. The number of data buses originating from the back-end of a controller typically corresponds to the number of ports available at its back-end. As can be seen from the figure, two of the buses on the back end of the local controller 120 connect directly to local storage devices 125. A collection of storage devices 125 at the back-end of the controller 120 may be considered a RAID. These disks 125 provide a medium for local storage of data to be used in conjunction with the operations of host computer 110 or other devices on the LAN.
For a variety of reasons, it may be desirable to store copies of main databases remotely as well as locally. Referring to the previous example, if a bank using a distributed computer system relies on its remote central databases for information and loses connectivity with the central databases by, for example, power outage or equipment failure, the overall system becomes inoperable. Similarly, if the central databases become corrupted through computer hacking or a computer virus, it may be undesirable to use the compromised databases. Therefore, a bank may wish to have several remote databases which are all kept current by, for example, the process of database shadowing. In such a system, should the bank lose the primary database site due to some loss of communications, database malfunction, or other such catastrophe, the bank has available another database site within its network containing the same information. The redundant site allows the bank to continue to transact business without an impact from the loss of their primary database site. Such a system is referred to as a "disaster tolerant" system.
Again referring to FIG. 1, one possible configuration of proprietary storage controllers used for storing data remotely and achieving a disaster tolerant system is shown. The storage controller is considered proprietary because the vendor of the controller does not make technical information about the interface 165 available to those who acquire or use the controller. In this example, a communication path 180 connects the proprietary interfaces of the controllers 120 and 140 to accomplish remote "out-of-band" communications between two compatible disk controllers. "Out-of-band" communications refers to data from host 110 that gets channeled off of the standard data bus within the local controller 120, which passes data from the front-end port to the back-end ports, and redirected to the proprietary interface 165. In such a proprietary disaster tolerant system, the communication path 180 comprises the following elements: local proprietary interface 165 within local controller 120, remote proprietary interface 175 within remote controller 140, local and remote data lines 160, 170, local and remote modems 135, 137 and long haul data link 136. The proprietary interfaces 165, 175 provide a physical and logical means by which to have data sent by the local superordinate controller 120 in a format which is interpretable by the remote subordinate controller 140. Once the data is received and interpreted by remote controller 140, it transmits the data to storage devices 145 at the back-end 150 of controller 140.
Because proprietary data is generally not made available to users, the disk controller vendor is often required to configure the proprietary interface within the disk controller for disaster tolerance. This typically results in a customized controller which is not fully maintainable by the owner of the system. Therefore, systems which use proprietary controllers to achieve remote database copies are relatively expensive in terms life cycle cost. "Life cycle cost" is the term used to encompass the expense of acquiring, configuring, operating, maintaining, and disposing of a device.

Method and apparatus for disaster tolerant computer system using cascaded storage controllers


A polypeptide substance isolate from rat serum which, upon administration to rats incapable of producing PTH (parathyroidectomized rats), produces an increase in the observed bone mineral apposition rate. The substance has been isolated in two forms, a first layer polypeptide having a molecular weight about twice that of a second smaller polypeptide. The first eleven amino acids of the sequence of the smaller polypeptide have been determined to be Gly Pro Gly Gly Ala Gly Glu Thr Lys Pro Ile (SEQ ID NO:3). The first seven amino acids of the larger polypeptide have been determined to be Gly Pro Gly Gly Ala Gly Glu (SEQ ID NO:2). The lager polypeptide might be the dimer of the smaller peptide. A nucleic acid probe, based on the amino acid sequence of the rat peptide was used to screen a human liver cDNA fetal library. A polypeptide was thus chemically synthesized according to the sequence Gly Ile Gly Lys Arg Thr Asn Glu His Thr Ala Asp Cys Lys Ile Lys Pro Asn Thr Leu His Lys Lys Ala Ala Glu Thr Leu Met Val Leu Asp Gln Asn Pro (SEQ ID NO:11). The bone apposition rate in rats increases in a dose dependent fashion upon administration of this chemically synthesized compound. A modified polypeptide containing a cysteinexe2x86x92alaine substinition was carried synthesized: Gly Ile Gly Lys Arg Thr Asn Glu His Thr Ala Asp Ala Lys Ile Lys Pro Asn Thr Leu His Lys Lys Ala Ala Glu Thr Met Val Leu Asp Gln Asn Gln Pro (SEQ ID NO:13). Some of the bone stimulatory effects of the normal polypeptide were found for the modified polypeptide.

The present invention relates to proteins and polypeptides which stimulate bone growth.
It is known that even in the adult human, bone can be subject to turnover. In certain locations, such as the internal auditory capsule, there is apparently no turnover after the organ is formed. In other locations, particularly in the central skeletal axis, the turnover appears to continue during adulthood. Bone turnover occurs on the surface of the existing bone matrix, which is composed of protein (mainly collagen) and minerals. Bone turnover is initiated with the destruction of bone matrix by osteoclasts. An osteoclast is a multinucleated cell which secretes acid and proteolytic enzymes leading to the lysis of the collagen matrix protein and the release of minerals into the extracelluar fluid compartment. Following this initial phase of bone destruction, or resorptive phase, formation of new bone protein matrix sets in. New bone proteins are deposited, and sometime later, minerals begin to be incorporated into the newly formed matrix. The formation of bone matrix and its subsequent mineralization are functions of osteoblasts, which are mononucleated cells. The formation phase is often followed by a period of inactivity (1,2). Resorption appears to be tightly coupled with formation (3) in vivo. Bone turnover is thus a succession of events, the location of which is known as the Bone Metabolism Unit or the BMU. Osteoblasts and osteoclasts, the putative mediators of bone turnover are thought to belong to two distinct cell lineages. These two cell types are not preformed cells, but they differentiate from their precursors through cell activation (4,5,6).
Bone matrix can either be maintained by a total cessation of bone turnover, as for the bone of the internal auditory capsule, or by a balance between formation and resorption. In many studies on skeletal changes in relation to age, a gain in the total body bone volume is observed during the growth period and the skeletal mass reaches a maximum at early adulthood. This gain is followed by a fall in bone volume as age advances. In females, a phase of more rapid bone loss often occurs during the perimenopausal period before a slower steadier phase. For this reason, bone loss in the female tends to be more severe than in the male. An understanding of bone balance in the BMU may thus be critical to understanding the pathogenesis of skeletal aging. In any case, mechanisms controlling bone turnover are complex and are not well understood at this time. The complexity of the control mechanisms has resulted in a variety of approaches to reducing bone loss.
Generally speaking, bone turnover can be regulated at two different stages. It can be regulated at the stage of the activation of precursor cells. Regulators of cellular activation can control not only the number of active BMU in the skeleton, but possibly also the number of osteoclasts and osteoblasts in an individual BMU. Alternatively, bone turnover can be regulated at the level of differentiated bone cells. The complexity of the bone cell system makes the separate study of these two levels of regulation difficult (3).
Regulators of bone cells appear to fall into two categories. The first of these interacts with specific receptors on cell membranes. One class of these regulators acts through the adenylate cyclase system with the generation of intra-cellular cyclic AMP as a second messenger acting on the protein kinase K system. Parathyroid hormone (PTH) and calcitonin (CT) belong to this class (7). A second class also interacts with a membrane receptor and results in the intracellular release of a molecule derived from phosphoinositides which in turn leads to an increase in intracellular calcium and activation of Kinase C. A third class involves interaction of the regulator with a cell surface receptor, but the second signal is generated by the receptor molecule itself with the subsequent activation of tyrosine Kinase. Many of the growth factors appear to act in this way (8-15). The second category of regulator does not interact with a cell membrane receptor, but can cross the cell membrane to bind with a cytosolic receptor. The regulator is then transported across the nuclear membrane by the cytosolic receptor to interact with the DNA resulting in increased transcription of specific genes. Steroid hormones, including vitamin D, appear to act in this manner (16).
Many hormones stimulate the proliferation of osteoclasts. These include 1,25(OH)2D, PTH and prostaglandins. PTH and 1,25(OH)2D receptors in osteoclasts have apparently not yet been identified. These two hormones seem to have no effect on osteoclasts in culture. However, when osteoclasts are co-cultured with osteoblast-like cell lines, PTH and 1,25(OH)2D stimulate the proliferation of osteoclasts. IL-1 and TNF appear to act in a similar way as PTH and 1,25(OH)2D. Other growth factors, like EGF, TFG and PDGF appear to stimulate osteoclasts through increased production of PGE. Calcitonin and corticosteroids are known osteoclast inhibitors along with chemicals such as diphosphonates.
It is currently believed that interleukin 1 may stimulate collagen and non-collagen bone protein and DNA synthesis. The effect on bone protein synthesis is blocked by indomethacin, suggesting that this action of IL-1 is mediated through PGE. Indomethacin seems to have no effect on the IL-1 effect on osteoblast DNA synthesis. In culture studies on osteoblast-like cell lines suggest that some locally produced growth factors stimulate DNA and collagen synthesis. In bone cell culture, PTH or Vitamin D suppresses collagen synthesis. This in vitro effect of PTH contrasts with the in vivo effect observed in human subjects and experimental animals. It has been demonstrated in rats and in human hyperparathyroid patients that PTH can stimulate the deposition of mineralized bone matrix. Preliminary clinical trial studies on the efficacy of the PTH 1-34 amino acid fragment in the treatment of osteoporosis indicate that this PTH fragment can increase the trabecular volume. The reason for this discrepancy is not yet fully explained.
Parathyroid hormone is a peptide of 84 amino acids in its mature form. Initially translated pre-pro-parathyroid hormone is much larger, the pre sequence being a signal sequence which is cleaved when the peptide enters the rough endoplasmic reticulum. In the golgi apparatus, the pro-sequence is cleaved off leaving the intact mature hormone packaged in the secretory granule. It appears that regulation of the rate of secretion is governed not so much by the rate of production of the intracellular peptide, but in the rate of intracellular destruction and in the rate of secretion. Intracellularly, the mature peptide is truncated at both the amino and the carboxyl termini. The truncated peptide may be secreted into the circulation as an inactive fragment. The secretion of the mature peptide can be stimulated by a drop in the extracelluar calcium concentration. An elevated serum calcium concentration on the other hand appears to suppress the secretion of PTH. Once in circulation, the mature peptide is rapidly cleaved in the liver at many sites of the molecule including the region of the 38 amino acid residue. The smaller fragment at the amino terminal end, which includes the first 34 amino acids, carries the full known biological activity in terms of its action on the kidney, the intestine and the bone. It also binds fully to the cell membrane receptor to stimulate cAMP production. The level of the 1-38 fragment in the serum is normally unmeasurable indicating that it has a short circulatory life. The larger inactive carboxyl terminal fragment has a relatively long half life and carries the highest proportion of the immunoreactive PTH in the circulatory system. All fragments in circulation are eventually destroyed in the kidney and the liver. One of the renal mechanisms for ridding the circulating inactive PTH fragments is glomerular filtration (17).
PTH participates in calcium and skeletal homeostasis. PTH stimulates the tubular resorption of calcium by the kidney and inhibits the reabsorption of phosphate and bicarbonate by the proximal renal tubules. A second effect of PTH on the kidney is the stimulation of 1,25(OH)2D production. This vitamin D metabolite is an in vivo stimulator of osteoclasts as well as an enhancer of intestinal calcium absorption. The increase in calcium absorption by the intestine following PTH stimulation is mediated by this vitamin D metabolite. In vivo, PTH stimulates osteoclastic bone resorption with the release of calcium into the circulation. PTH also causes proliferation of osteoblasts (18). In many cases of hyperparathyroidism there is a skeletal loss. However, an increase in spinal density has been reported in some cases of primary hyperparathyroidism (19,20,21) as well as in secondary hyperparathyroidism complicating renal failure. Kalu and Walker have observed that chronic administration of low doses of parathyroid extract led to sclerosis of bone in the rat (22). Tam et al. studied the effect of low calcium diet on the bone mineral apposition rate in the rat by tetracycline labelling and found that despite the loss of bone due to increase in bone resorption histologically (as a result of secondary hyperparathyroidism), the bone mineral apposition rate was increased (23). It was also found that the bone mineral apposition rate was increased in 23 human patients with mild primary hyperparathyroidism (24). After successful removal of parathyroid adenoma from four of the patients, the rate returned to the level observed in control subjects. There has also been found to be a dose dependent stimulation of the mineral apposition rate by PTH. The potency of the 1-34 fragment and the intact PTH hormone appears to be about the same on a molar basis. This is consistent with the 1-34 fragment of the PTH molecule carrying the biological activity of the intact hormone. It has also been observed that the end result of the administration of PTH on skeletal homeostasis depends on how the hormone is administered. For the same daily dose, the bone volume shows a dose dependent increase if the daily dose of the hormone is given as one single injection. However, when the same daily dose is administered by continuous infusion with a subcutaneous miniosmotic pump, the result is bone loss. Intermittent injection causes practically no effect on the serum calcium levels whereas infusion causes a dose dependent increase in the serum calcium. The effects of PTH administered by these two routes on bone mineral apposition rate as measured by tetracycline labelling are the same. What accounts for this differential effect is not understood (25).
Given the general understanding of bone growth and its regulation, various approaches to treatment of diseases involving reduction of bone mass and accompanying disorders are exemplified in the patent literature. For example, PCT Patent Application No. 9215615 published Sep. 17, 1992 describes a protein derived from a porcine pancreas which acts to depress serum calcium levels for treatment of bone disorders that cause elevation of serum calcium levels. European Patent Application No. 504938 published Sep. 23, 1992 describes the use of di- or tripeptides which inhibit cysteine protease in the treatment of bone diseases. PCT Patent Application No. 9214481 published Sep. 3, 1992 discloses a composition for inducing bone growth, the composition containing activin and bone morphogenic protein. European Patent Application No. 499242 published Aug. 19, 1992 describes the use of cell growth factor compositions thought to be useful in bone diseases involving bone mass reduction because they cause osteoblast proliferation. PCT Patent Application No. 4039656 published Jun. 17, 1992 describes a drug containing the human N-terminal PTH fragment 1-37. European Patent Application No. 451867 published Sep. 16, 1991 describes parathyroid hormone peptide antagonists for treating dysbolism associated with calcium or phosphoric acid, such as osteoporosis.
The relatively short half life of PTH in the blood serum and the relatively lengthy effect of intermittent PTH injection led the present investigator to the hypothesis that PTH may in some way lead to induction of a second factor into the circulatory system. The presence of such a second factor in blood serum of rats and of humans has thus been investigated.
It has been found possible to isolate from rat blood serum a polypeptide substance which, upon administration to rats incapable of producing PTH (parathyroidectomized rats), produces an increase in the observed bone mineral apposition rate. It has further been observed that the bone apposition rate increases with the dose of the isolated substance administered, at least over the dose range and time period studied. The substance has been isolated in two forms, a first larger polypeptide having a molecular weight about twice that of a second smaller polypeptide. The first eleven amino acids of the sequence of the smaller polypeptide have been determined to be Gly Pro Gly Gly Ala Gly Glu Thr Lys Pro Ile (SEQ ID NO:1). The first seven amino acids of the larger polypeptide have been determined to be Gly Pro Gly Gly Ala Gly Glu (SEQ ID NO:2). The similarity of these two NH2-terminal sequences has led to the proposition that the larger polypeptide might be the dimer of the first.
A nucleic acid probe, based on the amino acid sequence of the rat peptide has been synthesized and used to screen a human liver cDNA fetal library in order to isolate a human nucleic acid sequence coding for a human bone apposition polypeptide. A polypeptide was thus chemically synthesized according to the sequence Gly Ile Gly Lys Arg Thr Asn Glu His Thr Ala Asp Cys Lys Ile Lys Pro Asn Thr Leu His Lys Lys Ala Ala Glu Thr Leu Met Val Leu Asp Gln Asn Gln Pro (SEQ ID NO:11). It has been observed that the bone apposition rate in intact rats increases in a dose dependent fashion upon administration of this chemically synthesized compound. Reduced bone growth, normally observed for ovariectomized rats, was observed not to occur in rats after being administered with the polypeptide over a four week period beginning two weeks after ovariectomization. Bone calcium density was found to be maintained in ovariectomized rats administered with the polypeptide over an eight week period beginning eight weeks after ovariectomization.
It is thought possible that the active polypeptide is a dimer of the foregoing sequence, there being evidence of significant dimer formation, presumably due to a disulfide bridge between two polypeptides having the sequence shown.
A modified form of the polypeptide containing a cysxe2x86x92ala substitution was thus synthesized: Gly Ile Gly Lys Arg Thr Asn Glu His Thr Ala Asp Ala Lys Ile Lys Pro Asn Thr Leu His Lys Lys Ala Ala Glu Thr Leu Met Val Leu Asp Gln Asn Gln Pro (SEQ ID NO:13). Some of the bone stimulatory effects of the normal polypeptide were found for the modified polypeptide.
The bone mineral apposition rate in rats administered with rabbit anitbodies to the normal polypeptide (SEQ ID NO:11) was found to be suppressed. The suppression was found to be attenuated in rats administered with both the normal polypeptide and antibodies to same.